MassLynx NT Users Guide

MassLynx NT User’s Guide

Version 4.0
Waters Part No – 715000401

Micromass Part No – 6666661

30th October 2002

Issue 2

Page i
MassLynx NT Users Guide

MassLynx NT User’s Guide

The software described in this Manual is furnished under a license agreement and may be used
only in accordance with the terms of that agreement.

Copyright Notice

Micromass UK Limited believes that the information in this publication is accurate. However the
information is subject to change without notice and should not be construed as a contractual
undertaking by Micromass UK Limited. Despite the care that has been given to the preparation of
this publication, Micromass UK Limited accepts no responsibility for any loss or any other matter
that may arise from any error or inaccuracy that may inadvertently have been included.

Copyright  1993-2002 Micromass Ltd. All Rights Reserved.

No part of this publication may be copied without the express written permission of Micromass
UK Limited.

Trademarks

Micromass ® is a registered trademark of Micromass UK Limited (Reg. U.S. Pat. & Tm. Off.).

MassLynx and BioLynx are registered trademarks of Micromass UK Limited.

Windows is a trademark of Microsoft Corporation. Other product names mentioned in this manual
may be trademarks or registered trademarks of their respective companies and are hereby
acknowledged.

Page ii
 MassLynx NT Users Guide

Contents
MassLynx NT User’s Guide............................................................................................................. ii
Copyright Notice ................................................................................................................... ii
Trademarks............................................................................................................................ ii

Chapter 1 Introduction.................................................................................................................. 1-1

General .......................................................................................................................................... 1-3
Conventions................................................................................................................................... 1-3
New Features in MassLynx NT 4.0............................................................................................... 1-4
Installation.......................................................................................................................... 1-4
Inlets................................................................................................................................... 1-4
Instruments......................................................................................................................... 1-6
MassLynx General ............................................................................................................. 1-7
BioLynx ........................................................................................................................... 1-10
MetaboLynx ..................................................................................................................... 1-11
Molecule Viewer .............................................................................................................. 1-14
OpenLynx......................................................................................................................... 1-14
OpenLynx Global Server Version 1.0 (OLGS) ................................................................ 1-15
ProteinLynx...................................................................................................................... 1-17
ProteinLynx Global Server V1.1...................................................................................... 1-17
QuanLynx......................................................................................................................... 1-18
QuanOptimize .................................................................................................................. 1-19
New Features in MassLynx NT 3.5............................................................................................. 1-22
Supported Operating Systems .......................................................................................... 1-22
LCT .................................................................................................................................. 1-22
QTof ................................................................................................................................. 1-22
Quattro Ultima (EPCAS) ................................................................................................. 1-23
ZQ .................................................................................................................................... 1-23
AutoSpec NT.................................................................................................................... 1-23
GCT.................................................................................................................................. 1-23
Waters .............................................................................................................................. 1-23
Gilson ............................................................................................................................... 1-23
Shutdown ......................................................................................................................... 1-23
HP6890............................................................................................................................. 1-23
Jasco ................................................................................................................................. 1-24
CTC PAL ......................................................................................................................... 1-24
MassLynx General ........................................................................................................... 1-24
DataBridge ....................................................................................................................... 1-24
Quantify ........................................................................................................................... 1-24
OpenLynx......................................................................................................................... 1-24
MetaboLynx ..................................................................................................................... 1-25
FractionLynx .................................................................................................................... 1-26
MicrobeLynx.................................................................................................................... 1-26
ProteinLynx Global Server 1.0......................................................................................... 1-26
Web browser based database searching ........................................................................... 1-27
BioLynx ........................................................................................................................... 1-27
ProteinLynx...................................................................................................................... 1-27
ProteinProbe..................................................................................................................... 1-27
New Features in MassLynx NT 3.4............................................................................................. 1-28
MassLynx......................................................................................................................... 1-28
BioLynx ........................................................................................................................... 1-28
ProteinLynx...................................................................................................................... 1-28
OpenLynx......................................................................................................................... 1-28
NeoLynx........................................................................................................................... 1-29
MetaboLynx ..................................................................................................................... 1-29
FractionLynx .................................................................................................................... 1-29
All File Accurate Mass Measure ...................................................................................... 1-29
Combine All Files ............................................................................................................ 1-29

Page iii
MassLynx NT Users Guide

Calibration........................................................................................................................ 1-29
Waters CapLC.................................................................................................................. 1-29
Waters 2700 ..................................................................................................................... 1-29
Jasco Systems................................................................................................................... 1-29
CTC PAL Autosampler.................................................................................................... 1-30
Support Removed............................................................................................................. 1-30
Embedded PC Support ..................................................................................................... 1-30
AutoSpec.......................................................................................................................... 1-30
QTof................................................................................................................................. 1-30
LCT .................................................................................................................................. 1-30
MALDI-Tof ..................................................................................................................... 1-30
IsoPrime ........................................................................................................................... 1-31
Platform ICP .................................................................................................................... 1-31
New Features in MassLynx NT 3.3............................................................................................. 1-31
BioLynx ........................................................................................................................... 1-31
ProteinLynx...................................................................................................................... 1-31
AutoSpec Support ............................................................................................................ 1-31
LCT .................................................................................................................................. 1-31
MetaboLynx ..................................................................................................................... 1-31
QuanLynx ........................................................................................................................ 1-32
Elemental ......................................................................................................................... 1-32
Map .................................................................................................................................. 1-32
Gilson Pumps ................................................................................................................... 1-32
Autosamplers ................................................................................................................... 1-32
Strip.................................................................................................................................. 1-32
Quantify ........................................................................................................................... 1-32
Spectrum .......................................................................................................................... 1-32
OpenLynx ........................................................................................................................ 1-32
Automatic Instrument Shutdown ..................................................................................... 1-33
Bio-Q/Quattro Family ...................................................................................................... 1-33
Example Macros .............................................................................................................. 1-33
Divert Valve support for Platform LCZ/ZMD and Quattro LC ....................................... 1-33
New Features in MassLynx NT 3.2............................................................................................. 1-33
BioLynx ........................................................................................................................... 1-33
ProteinLynx...................................................................................................................... 1-34
MaxEnt 3 .......................................................................................................................... 1-34
OpenLynx ........................................................................................................................ 1-34
Support for Waters 2700 Autosampler............................................................................. 1-34
Support for Waters 486 UV Detector............................................................................... 1-34
Support for Waters 2487 UV Detector............................................................................. 1-34
Support for Waters 600 Pump.......................................................................................... 1-34
Accurate Mass Chromatograms ....................................................................................... 1-34
AutoLynx ......................................................................................................................... 1-34
All File Accurate Mass Measure...................................................................................... 1-35
Database Logging ............................................................................................................ 1-35
Elemental Composition.................................................................................................... 1-35
Quantify ........................................................................................................................... 1-35
Scan Function Editor........................................................................................................ 1-35
Priority Processing ........................................................................................................... 1-35
Night Time Processing..................................................................................................... 1-35
MassLynx Status File....................................................................................................... 1-35
Import Worksheet ............................................................................................................ 1-35
New Features in MassLynx NT 3.1............................................................................................. 1-35
BioLynx ........................................................................................................................... 1-36
OpenLynx ........................................................................................................................ 1-36
Support for Waters 2690 LC Systems via GPIB interface ............................................... 1-36
Support for Waters 996 PDA Detector............................................................................. 1-36
New Features in MassLynx NT 3.0............................................................................................. 1-36
MassLynx top level screen............................................................................................... 1-36
Sample Lists..................................................................................................................... 1-36

Page iv
 MassLynx NT Users Guide

Project Wizard.................................................................................................................. 1-36

Projects............................................................................................................................. 1-36
Quantify ........................................................................................................................... 1-37
Longer File Names ........................................................................................................... 1-37
Analytical Component Engine ......................................................................................... 1-37
OpenLynx......................................................................................................................... 1-37
Q-Tof................................................................................................................................ 1-37
Chromatogram Peak Purity .............................................................................................. 1-37
Chromatogram Signal to Noise ........................................................................................ 1-37
Chromatogram Retention Index ....................................................................................... 1-37
Support for Gilson Pumps ................................................................................................ 1-37
Support for HP6890 GC Systems..................................................................................... 1-37
Support for Jasco LC Systems.......................................................................................... 1-38
Support for Waters 2690 LC Systems .............................................................................. 1-38
Support for CTC/LEAP Technology PAL autosampler................................................... 1-38
Support for MicroTech LC Systems................................................................................. 1-38
Periodic Table .................................................................................................................. 1-38
BioLynx ........................................................................................................................... 1-38
Colors and Fonts .............................................................................................................. 1-38
Startup and Shutdown ...................................................................................................... 1-38
Macro support .................................................................................................................. 1-38

Chapter 2 Installing MassLynx..................................................................................................... 2-1

Minimum System Requirements ................................................................................................... 2-3
Installing MassLynx ...................................................................................................................... 2-4
Preparation ......................................................................................................................... 2-4
To Install MassLynx........................................................................................................... 2-4
Using the MassLynx Installation Wizard ...................................................................................... 2-5
General ............................................................................................................................... 2-5
Tool Tips ............................................................................................................................ 2-5
The MassLynx Security Options and Backup Scheduler Dialogs...................................... 2-5
The MassLynx Options, MassLynx Installation directory dialog ...................................... 2-6
Failed Installation, or Installation Cancelled...................................................................... 2-7
The MassLynx Folder.................................................................................................................... 2-7
Installing Updates to MassLynx .................................................................................................... 2-8
Installing Additional MassLynx Application Managers................................................................ 2-8
General ............................................................................................................................... 2-8
Procedure ........................................................................................................................... 2-8
Installing MassLynx Libraries....................................................................................................... 2-9
Installing the SWISS-PROT Database ........................................................................................ 2-10
Uninstalling MassLynx ............................................................................................................... 2-10
Repairing MassLynx ................................................................................................................... 2-11

Chapter 3 The MassLynx Window and Related Information..................................................... 3-1

Opening MassLynx ....................................................................................................................... 3-5
Closing MassLynx......................................................................................................................... 3-5
The MassLynx Window ................................................................................................................ 3-6
The MassLynx Menu Bar .............................................................................................................. 3-7
General ............................................................................................................................... 3-7
The File Menu .................................................................................................................... 3-7
The View Menu.................................................................................................................. 3-9
The Run Menu.................................................................................................................... 3-9
The Security Menu ............................................................................................................. 3-9
The Help Menu ................................................................................................................ 3-10
The MassLynx Tool Bar.............................................................................................................. 3-10
General ............................................................................................................................. 3-10
The Sample List Menu Bar.......................................................................................................... 3-11
The Sample List........................................................................................................................... 3-11
The MassLynx Bar ...................................................................................................................... 3-11
General ............................................................................................................................. 3-11

Page v
MassLynx NT Users Guide

Displaying the MassLynx Bar.......................................................................................... 3-12

The Shortcut Bar .............................................................................................................. 3-12
The Queue Bar ................................................................................................................. 3-16
The MS Status Bar ........................................................................................................... 3-17
Changing the Colors and Fonts in MassLynx Windows ............................................................. 3-18
The Colors and Fonts dialog ............................................................................................ 3-18
The Font Dialog ............................................................................................................... 3-19
The Color Dialog ............................................................................................................. 3-20
The Define Custom Color Dialog .................................................................................... 3-21
To Change MassLynx Fonts or Colors............................................................................. 3-21
MassLynx System Global Parameters ......................................................................................... 3-22
General............................................................................................................................. 3-22
The Options Dialog .......................................................................................................... 3-22
The Options, Multi-probe Dialog..................................................................................... 3-25
The Masslynx.ini File.................................................................................................................. 3-26
General............................................................................................................................. 3-26
To Restore Masslynx.sav Using the Windows Explorer.................................................. 3-26
Selecting and Viewing Data ........................................................................................................ 3-27
General............................................................................................................................. 3-27
Opening Data Files: The MassLynx Window Data Browser Dialog ............................... 3-27
The History Selector Dialog............................................................................................. 3-28
The Experimental Record Window.................................................................................. 3-31
Using Windows Explorer to Open Multiple Data Files............................................................... 3-32
Projects ........................................................................................................................................ 3-33
General............................................................................................................................. 3-33
To Create a New Project .................................................................................................. 3-33
To Open an Existing Project ............................................................................................ 3-35
Directory Structure ...................................................................................................................... 3-35
Data File Structure ........................................................................................................... 3-37
Displaying Spectra ...................................................................................................................... 3-37
To Display a Spectrum Using the Sample List Menu Bar ............................................... 3-37
To Display a Spectrum from a Chromatogram ................................................................ 3-37
To Remove Spectra and Spectrum Windows................................................................... 3-38
Displaying Chromatograms......................................................................................................... 3-38
To Display a Chromatogram Using the Sample List Menu Bar....................................... 3-38
To Display a Chromatogram from Spectrum ................................................................... 3-38
To Remove Chromatogram Traces and Chromatogram Windows .................................. 3-38
The Header Editor Dialog ........................................................................................................... 3-39
General............................................................................................................................. 3-39
The User Text Dialog....................................................................................................... 3-40
The Format Dialog ........................................................................................................... 3-41
To Add Information to the Displayed Header in a MassLynx Program Window ............ 3-41
To Remove Information from the Displayed Header....................................................... 3-42
Printing Data ............................................................................................................................... 3-42
General............................................................................................................................. 3-42
Printing a Specific MassLynx Window Using the Tool Bar ............................................ 3-43
Printing a Specific MassLynx Window Using the Menu Bar .......................................... 3-43
Window Commands .................................................................................................................... 3-44
Getting Help ................................................................................................................................ 3-44

Chapter 4 Sample Lists .................................................................................................................4-1

Introduction ................................................................................................................................... 4-5
The Sample List Menu Bar............................................................................................................ 4-6
General............................................................................................................................... 4-6
Commands ......................................................................................................................... 4-6
The Sample List Menu Bar Edit Menu .............................................................................. 4-6
The Sample List Menu Bar Samples Menu........................................................................ 4-7
Pop-up Menus ............................................................................................................................. 4-10
General............................................................................................................................. 4-10
Pop-up Menu Commands Not Available on the Sample List Menu Bar ......................... 4-11

Page vi
 MassLynx NT Users Guide

Opening an Existing Sample List ................................................................................................ 4-11

Saving a Sample List ................................................................................................................... 4-12
Printing Sample Lists .................................................................................................................. 4-13
Creating Sample Lists.................................................................................................................. 4-13
Creating a Sample List in the MassLynx Window........................................................... 4-13
Creating a Sample List by Copying a Spreadsheet........................................................... 4-13
Importing a Worksheet into the Sample List Editor .................................................................... 4-14
General ............................................................................................................................. 4-14
To Import a Worksheet .................................................................................................... 4-14
Creating Import Worksheet Files ..................................................................................... 4-15
Access .............................................................................................................................. 4-16
Excel................................................................................................................................. 4-17
Notepad ............................................................................................................................ 4-17
Importing Data ............................................................................................................................ 4-17
General ............................................................................................................................. 4-17
To Import Data ................................................................................................................. 4-18
Creating Import Data Files ............................................................................................... 4-18
Access .............................................................................................................................. 4-18
Excel................................................................................................................................. 4-19
Notepad ............................................................................................................................ 4-19
Sample List Formats.................................................................................................................... 4-19
General ............................................................................................................................. 4-19
Loading an Existing Sample List Format ......................................................................... 4-20
Saving a Sample List Format ........................................................................................... 4-20
Changing the Sample List Format.................................................................................... 4-21
Manipulating the Sample List Editor Display ............................................................................. 4-23
Selecting Areas of the Display ......................................................................................... 4-23
Editing Data in Fields .................................................................................................................. 4-24
General ............................................................................................................................. 4-24
Fields Containing Files..................................................................................................... 4-24
Fields with Multiple Options............................................................................................ 4-25
Directly-Editable Cells..................................................................................................... 4-26
Editing Data in an Area ............................................................................................................... 4-26
Inserting Rows in a Sample List.................................................................................................. 4-26
Adding Samples to the Bottom of a Sample List......................................................................... 4-27
Cutting Data ................................................................................................................................ 4-27
Cutting Data from a Directly-Editable Cell...................................................................... 4-27
Cutting the Contents of Multiple Cells............................................................................. 4-27
Cutting Samples from the Sample List............................................................................. 4-27
Deleting Data............................................................................................................................... 4-28
Deleting Data from Cells.................................................................................................. 4-28
Deleting Data from Columns ........................................................................................... 4-28
Deleting Rows.................................................................................................................. 4-28
Deleting Data from the Whole Sample List ..................................................................... 4-29
Pasting Data................................................................................................................................. 4-29
Pasting Data into a Directly-Editable Cell ....................................................................... 4-29
Pasting into Multiple Cells ............................................................................................... 4-29
Pasting Samples into the SLE .......................................................................................... 4-29
Sorting the Rows in a Sample List .............................................................................................. 4-30
Specifying the Number of Injections for Each Sample ............................................................... 4-30
Action On Error........................................................................................................................... 4-31
Updating the Sample List from the AutoSampler Bed Layout.................................................... 4-32
The AutoSampler Bed Layout Dialog.............................................................................. 4-32

Chapter 5 The MassLynx Queue.................................................................................................. 5-1

Introduction ................................................................................................................................... 5-3
Starting Data Acquisition .............................................................................................................. 5-3
The Queue Bar............................................................................................................................... 5-5
General ............................................................................................................................... 5-5
Changing the Job Properties............................................................................................... 5-6

Page vii
MassLynx NT Users Guide

Changing the Queue Properties.......................................................................................... 5-7

Chapter 6 Chromatogram .............................................................................................................6-1

Getting Started............................................................................................................................... 6-7
To Display the Total Ion Current (TIC) Chromatogram .................................................... 6-7
To Display a Summed Mass Chromatogram Around a Peak in a Spectrum...................... 6-7
The Chromatogram Window......................................................................................................... 6-7
The Chromatogram Menu Bar....................................................................................................... 6-8
The Chromatogram File Menu........................................................................................... 6-8
The Chromatogram Edit Menu .......................................................................................... 6-8
The Chromatogram Display Menu..................................................................................... 6-9
The Chromatogram Process Menu ................................................................................... 6-12
The Chromatogram Window Menu ................................................................................. 6-13
The Chromatogram Tools Menu ...................................................................................... 6-14
The Chromatogram Help Menu ....................................................................................... 6-14
The Chromatogram Tool Bar ...................................................................................................... 6-15
General............................................................................................................................. 6-15
Customizing the Chromatogram Tool Bar ....................................................................... 6-16
Displaying Chromatograms......................................................................................................... 6-19
Adding or Replacing Chromatogram Traces.................................................................... 6-19
The New Chromatogram Dialog ...................................................................................... 6-20
The Mass Chromatogram dialog ...................................................................................... 6-20
TIC and BPI Chromatograms........................................................................................... 6-24
Analog Chromatograms ................................................................................................... 6-25
Aligning Analog Chromatograms .................................................................................... 6-26
The Chromatogram Pointer ......................................................................................................... 6-27
Manipulating the Display ............................................................................................................ 6-27
Altering the Range of the Horizontal Axis....................................................................... 6-27
Centering the Display Around a Particular Point ............................................................. 6-28
Altering the Range of the Intensity Axis.......................................................................... 6-29
Altering the Range of Both Axes ..................................................................................... 6-29
Setting Magnified Ranges ................................................................................................ 6-29
Deleting Magnification Ranges........................................................................................ 6-30
Restoring the Display....................................................................................................... 6-31
Setting the Display Range Defaults ................................................................................. 6-31
Controlling the Appearance of the Display ................................................................................. 6-32
General............................................................................................................................. 6-32
To Change the Display Parameters .................................................................................. 6-32
Controlling the Appearance of Peak Labels ................................................................................ 6-34
General............................................................................................................................. 6-34
To Change the Peak Annotation Parameters .................................................................... 6-34
Removing Chromatograms from the Display.............................................................................. 6-36
To Remove a Single Chromatogram Trace from the Display .......................................... 6-36
To Remove Multiple Chromatogram Traces from the Display........................................ 6-36
Real-Time Display of Chromatograms........................................................................................ 6-36
Changing the Order of Displayed Chromatograms ..................................................................... 6-37
Adding Text to the Chromatogram Display ................................................................................ 6-37
Processing Chromatograms ......................................................................................................... 6-38
General............................................................................................................................. 6-38
Processing Multiple Chromatograms ............................................................................... 6-38
Background Subtract ................................................................................................................... 6-39
General............................................................................................................................. 6-39
Performing a Background Subtract .................................................................................. 6-39
Example of Background Subtract .................................................................................... 6-40
Checking the Results of Background Subtract................................................................. 6-41
Smoothing Chromatograms......................................................................................................... 6-41
General............................................................................................................................. 6-41
The Smooth Chromatogram Dialog ................................................................................. 6-41
To Smooth a Chromatogram ............................................................................................ 6-42
Integrating Chromatograms......................................................................................................... 6-43

Page viii
 MassLynx NT Users Guide

General ............................................................................................................................. 6-43

To Integrate a Chromatogram .......................................................................................... 6-44
Standard Peak Detection Parameters................................................................................ 6-47
ApexTrack Peak Detection Parameters ............................................................................ 6-52
Peak Thresholding............................................................................................................ 6-53
To Display Information about an Integrated Peak............................................................ 6-54
Editing Integrated Peaks................................................................................................... 6-54
Peak Purity .................................................................................................................................. 6-57
General ............................................................................................................................. 6-57
To Calculate the Peak Purity Index for a Total Ion Chromatogram................................. 6-59
Signal to Noise Ratio................................................................................................................... 6-59
General ............................................................................................................................. 6-59
The Signal To Noise dialog.............................................................................................. 6-60
To Calculate the Signal to Noise Value for a Mass Chromatogram................................. 6-61
Combine Spectra ......................................................................................................................... 6-61
ElectroSpray Data Processing – Components ............................................................................. 6-62
General ............................................................................................................................. 6-62
Component Identification................................................................................................. 6-62
The Auto Find Components Dialog ................................................................................. 6-63
Automatic Component Finding ........................................................................................ 6-64
The Component Worklist Dialog ..................................................................................... 6-66
Peak Lists .................................................................................................................................... 6-69
General ............................................................................................................................. 6-69
The Edit Peak List Dialog ................................................................................................ 6-69
To Create a New Peak List File........................................................................................ 6-71
To Open an Existing a Peak List File............................................................................... 6-72
To Append Peaks to the Current Peak List ...................................................................... 6-72
To Delete Peaks from the Current Peak List .................................................................... 6-72
Reading a Peak List into a Chromatogram....................................................................... 6-72
Chromatogram Display When Switching Between MS and MS/MS Modes .............................. 6-74
Copying To and From the Windows Clipboard........................................................................... 6-75
General ............................................................................................................................. 6-75
To Copy a Chromatogram as a Picture to the Clipboard.................................................. 6-75
To Copy a Chromatogram as a Text List to the Clipboard............................................... 6-75
To Copy Integrated Chromatogram Peaks as a Text List to the Clipboard ...................... 6-75
To Paste Information from the Windows Clipboard into a Chromatogram Window....... 6-75
Removing Pasted Input from the Display ........................................................................ 6-76
Retention Index ........................................................................................................................... 6-76
General ............................................................................................................................. 6-76
The Retention Index Table dialog .................................................................................... 6-76
To Set Up the Retention Index Table ............................................................................... 6-77
To Delete the Retention Index Table................................................................................ 6-77
To Make a Retention Index Calibration ........................................................................... 6-77
To Display Retention Index Values on a Chromatogram................................................. 6-78
To Check Retention Index Calibration Status .................................................................. 6-79

Chapter 7 Spectrum ...................................................................................................................... 7-1

Getting Started............................................................................................................................... 7-7
To Display the First Scan of the Current Data File ............................................................ 7-7
To Display a Scan at a Particular Time in the Current Data File ....................................... 7-7
The Spectrum Window.................................................................................................................. 7-8
The Spectrum Menu Bar................................................................................................................ 7-9
The Spectrum File Menu.................................................................................................... 7-9
The Spectrum Edit Menu ................................................................................................... 7-9
The Spectrum Display Menu............................................................................................ 7-10
The Spectrum Process Menu............................................................................................ 7-13
The Spectrum Tools Menu ............................................................................................... 7-15
The Spectrum Window Menu .......................................................................................... 7-16
The Spectrum Help Menu ................................................................................................ 7-17
The Spectrum Tool Bar ............................................................................................................... 7-17

Page ix
MassLynx NT Users Guide

General............................................................................................................................. 7-17
Customizing the Spectrum Tool Bar ................................................................................ 7-18
Displaying Spectra ...................................................................................................................... 7-21
Adding or Replacing Spectra ........................................................................................... 7-21
The New Spectrum Dialog............................................................................................... 7-22
Viewing a Peak List Entry ............................................................................................... 7-22
Manipulating the Display ............................................................................................................ 7-23
Altering the Range of the Mass Axis ............................................................................... 7-23
Altering the Range of the Intensity Axis.......................................................................... 7-23
Altering the Range of Both Axes ..................................................................................... 7-23
Setting Magnified Ranges ................................................................................................ 7-23
Deleting Magnification Ranges........................................................................................ 7-25
Restoring the Display....................................................................................................... 7-25
Setting the Display Range Defaults ................................................................................. 7-26
Displaying a Spectrum as a List....................................................................................... 7-26
Controlling the Appearance of the Display ................................................................................. 7-28
General............................................................................................................................. 7-28
To Change the Display Parameters .................................................................................. 7-28
Controlling the Appearance of Peak Labels ................................................................................ 7-31
General............................................................................................................................. 7-31
To Change the Peak Annotation Parameters .................................................................... 7-31
To Annotate a Particular Peak.......................................................................................... 7-33
Removing Spectra from the Display ........................................................................................... 7-33
To Remove a Single Spectrum Trace from the Display................................................... 7-33
To Remove Multiple Spectrum Traces from the Display................................................. 7-33
Real-Time Display of Spectra .......................................................................................... 7-33
Changing the Order of Displayed Spectra........................................................................ 7-34
Adding Text to the Spectrum Display.............................................................................. 7-34
Exporting SEQUEST Files.......................................................................................................... 7-34
General............................................................................................................................. 7-34
To Export a SEQUEST File ............................................................................................. 7-35
Processing Spectra....................................................................................................................... 7-35
General............................................................................................................................. 7-35
Saving and Recalling Processed Spectra.......................................................................... 7-36
The Refine Process...................................................................................................................... 7-38
General............................................................................................................................. 7-38
To Refine a Scan in a Centroid-Mode Data File .............................................................. 7-38
The Combine Spectra Process ..................................................................................................... 7-39
General............................................................................................................................. 7-39
To Combine Scans in a Centroid-Mode Data File ........................................................... 7-39
The Background Subtract Process............................................................................................... 7-40
General............................................................................................................................. 7-40
To Subtract the Background from a Continuum Spectrum .............................................. 7-40
The Smooth Process .................................................................................................................... 7-41
General............................................................................................................................. 7-41
To Smooth a Continuum Spectrum.................................................................................. 7-42
The Center Process...................................................................................................................... 7-43
General............................................................................................................................. 7-43
To Center a Continuum Spectrum.................................................................................... 7-44
The Mass Measure Process ......................................................................................................... 7-46
General............................................................................................................................. 7-46
QTOF Accurate Mass ...................................................................................................... 7-47
The TOF Transform Process ....................................................................................................... 7-47
The Integration Process ............................................................................................................... 7-48
General............................................................................................................................. 7-48
To Integrate a Spectrum ................................................................................................... 7-48
ElectroSpray Data Processing ..................................................................................................... 7-49
General............................................................................................................................. 7-49
Setting Adduct Mass for Transform and MaxEnt ............................................................ 7-50
Finding Components for Transform................................................................................. 7-50

Page x
 MassLynx NT Users Guide

Editing Components for Transform.................................................................................. 7-54

The Transform Process..................................................................................................... 7-58
MaxEnt 1.......................................................................................................................... 7-59
MaxEnt Errors.................................................................................................................. 7-68
MaxEnt 2.......................................................................................................................... 7-69
MaxEnt 3.......................................................................................................................... 7-71
Isotope Cluster Abundance Plots................................................................................................. 7-75
General ............................................................................................................................. 7-75
The Isotope modelling Dialog.......................................................................................... 7-75
The User-definable elements Dialog ................................................................................ 7-77
To Produce an Isotope Cluster Abundance Plot............................................................... 7-78
Elemental Composition ............................................................................................................... 7-78
General ............................................................................................................................. 7-78
The EleComp Parameters Dialog ..................................................................................... 7-78
To Produce an Elemental Composition Report ................................................................ 7-79
To Update an Elemental Composition Report.................................................................. 7-80
The Elemental Composition Window .............................................................................. 7-80
The Elemental Composition Window Menu Bar ............................................................. 7-82
The Elemental Composition Window Tool Bar ............................................................... 7-84
Elemental Composition Parameters............................................................................................. 7-84
General ............................................................................................................................. 7-84
The Parameters Dialog; General Parameters Page ........................................................... 7-85
The Parameters Dialog; Symbol Parameters Page ........................................................... 7-87
Superatom Tables............................................................................................................. 7-91
Superatom Limits ............................................................................................................. 7-93
Performing a Calibration ............................................................................................................. 7-94
General ............................................................................................................................. 7-94
To Make a New Calibration ............................................................................................. 7-94
To Apply a Calibration..................................................................................................... 7-96
To Modify a Calibration................................................................................................... 7-97
Lock Mass ........................................................................................................................ 7-97
Copying to and from the Windows Clipboard............................................................................. 7-97
General ............................................................................................................................. 7-97
To Copy a Spectrum as a Picture to the Clipboard........................................................... 7-98
To Copy a Spectrum as a Text List to the Clipboard ....................................................... 7-98
To Paste Information from the Windows Clipboard into a Spectrum Window ............... 7-98
Removing Pasted Input from the Display ........................................................................ 7-98
Manipulating Library Spectra...................................................................................................... 7-99
To Display a Library Entry .............................................................................................. 7-99
To Append the Current Spectrum to the Current Library................................................. 7-99

Chapter 8 Strip and Combine Functions .................................................................................... 8-1

The Strip Program ......................................................................................................................... 8-5
General ............................................................................................................................... 8-5
The Strip Datafile Dialog ................................................................................................... 8-6
The Strip Datafile Dialog Menu Bar .................................................................................. 8-7
Creating an Enhanced Data File ......................................................................................... 8-8
Creating a Subtracted Data File........................................................................................ 8-10
Creating a Clustered Data File ......................................................................................... 8-11
Creating a CODA Data File ............................................................................................. 8-12
Selecting an Input Data File to Process............................................................................ 8-13
Selecting a Sub-range to Process...................................................................................... 8-14
Selecting a Background Data File .................................................................................... 8-15
Selecting an Output Data File .......................................................................................... 8-16
Setting Subtract Datafile Options..................................................................................... 8-16
Setting Enhance Datafile Options .................................................................................... 8-17
Setting Cluster Datafile Options....................................................................................... 8-18
Setting Cluster Centroid Options ..................................................................................... 8-19
Setting CODA Options..................................................................................................... 8-21
Stopping a Process ........................................................................................................... 8-21

Page xi
MassLynx NT Users Guide

The Combine Functions Program................................................................................................ 8-21

General............................................................................................................................. 8-21
Creating a Combined Data File........................................................................................ 8-22
The Combine Datafile Functions Dialog Menu Bar......................................................... 8-23
Selecting an Input Data File ............................................................................................. 8-24
Selecting a Sub-range to Process ..................................................................................... 8-25
Selecting an Output Data File .......................................................................................... 8-26
Stopping a Process ........................................................................................................... 8-26
The Combine All Files Program.................................................................................................. 8-26
General............................................................................................................................. 8-26
The Combine All Files Dialog Input File(s) List ............................................................. 8-28
The Combine All Files Dialog Menu Bar ........................................................................ 8-29
Processing Files................................................................................................................ 8-29

Chapter 9 Map ................................................................................................................................9-1

Introduction ................................................................................................................................... 9-3
Creating a Data File Map .............................................................................................................. 9-4
To Create a Data File Map ................................................................................................. 9-4
To Stop the Map Process Before it has been Completed ................................................... 9-5
The Map Tool Bar ......................................................................................................................... 9-6
General............................................................................................................................... 9-6
Selecting a Range to Map from the Data File................................................................................ 9-6
Manipulating the Display .............................................................................................................. 9-7
Altering the Display Range with the Mouse ...................................................................... 9-7
Altering the Display Range from the Menu ....................................................................... 9-8
To Change the Map Intensity Scaling ................................................................................ 9-8
Controlling the Appearance of the Display ........................................................................ 9-9
Displaying Diode Array Data........................................................................................... 9-10
Aligning Diode Array Data .............................................................................................. 9-10
Displaying the Chromatogram and Spectrum Windows .................................................. 9-11
The Status Bar.................................................................................................................. 9-11
Selecting the Current Cursor Position .............................................................................. 9-11
User Defined Cursor Positions......................................................................................... 9-13
Editing the Header Information........................................................................................ 9-13
Printing from Map ....................................................................................................................... 9-13
Copying to the Windows Clipboard ............................................................................................ 9-13
General............................................................................................................................. 9-13

Chapter 10 Quantify.....................................................................................................................10-1
Introduction ................................................................................................................................. 10-5
Accessing Quantify ..................................................................................................................... 10-5
MassLynx Automated Quantification - an Overview.................................................................. 10-6
How Does MassLynx Quantify and Report a List of Samples? .................................................. 10-7
Integration of Chromatograms ......................................................................................... 10-7
Generation of Calibration Curves .................................................................................... 10-8
Calculation of Compound Concentrations ....................................................................... 10-9
Displaying Quantify Results .......................................................................................... 10-10
A Step by Step Guide to Quantification .................................................................................... 10-10
1. Create a Sample List ................................................................................................. 10-10
2. Create a Quantify Method ......................................................................................... 10-11
3. Start the Analysis ...................................................................................................... 10-20
4. Quantify the Data ...................................................................................................... 10-22
Using the Quantify Window to Examine Results...................................................................... 10-23
General........................................................................................................................... 10-23
The Quantify Window Tool Bar .................................................................................... 10-23
Controlling the Contents of the Quantify Window ........................................................ 10-24
The Quantify Summary Window ................................................................................... 10-25
Selecting the Fields to be Displayed in the Summary Window and Summary Reports. 10-26
To Save the Summary Window ..................................................................................... 10-28
The Quantify Graphs Window ....................................................................................... 10-28

Page xii
 MassLynx NT Users Guide

The Quantify Peak List Window.................................................................................... 10-30

Manually Changing Quantify Results ....................................................................................... 10-32
General ........................................................................................................................... 10-32
Manual Peak Integration ................................................................................................ 10-32
To Exclude Erroneous Calibration Points ...................................................................... 10-34
To Prevent a Complete Sample from being Used to Create the Calibration Curve........ 10-35
To Perform Any of the Quantify Processes.................................................................... 10-35
Printing Quantify Reports.......................................................................................................... 10-36
General ........................................................................................................................... 10-36
Changing the Printed Quantify Reports Format ............................................................. 10-37
Selecting the Chromatogram Display Range for the Quantify Sample Report.......................... 10-39
Copying Quantify Summary Reports to the Clipboard.............................................................. 10-39
Exporting to a LIMS File........................................................................................................... 10-39
General ........................................................................................................................... 10-39
The Header Section ........................................................................................................ 10-40
The Samples Section ...................................................................................................... 10-40
The Calibration Section.................................................................................................. 10-41
Files Used During Quantify....................................................................................................... 10-41
General ........................................................................................................................... 10-41
The Sample List (.SPL) File........................................................................................... 10-41
The Quantify Method (.MDB) File ................................................................................ 10-42
Peak Lists (.PDB) File.................................................................................................... 10-42
Calibration Curves (.CDB) File...................................................................................... 10-42

Chapter 11 Library....................................................................................................................... 11-1

Introduction ................................................................................................................................. 11-5
The Library Tool Bar................................................................................................................... 11-6
Library Searching ........................................................................................................................ 11-6
General ............................................................................................................................. 11-6
The Presearch ................................................................................................................... 11-6
The Mainsearch................................................................................................................ 11-7
An Overview of Library Searching .................................................................................. 11-7
Selecting Which Libraries to Search ................................................................................ 11-8
Selecting a New Search Spectrum.................................................................................. 11-10
Changing the Library Search Parameters ....................................................................... 11-11
Library Search Filters..................................................................................................... 11-13
Starting a Library Search................................................................................................ 11-15
Automatic Library Search .............................................................................................. 11-15
Library Search Results .............................................................................................................. 11-16
General ........................................................................................................................... 11-16
Manipulating the Library Display .................................................................................. 11-17
The Hit List Window................................................................................................................. 11-18
General ........................................................................................................................... 11-18
Formatting the Hit List................................................................................................... 11-19
The Hits Window ...................................................................................................................... 11-21
General ........................................................................................................................... 11-21
Manipulating the Hits Window Display......................................................................... 11-21
The Delta Window .................................................................................................................... 11-22
The Structure Window .............................................................................................................. 11-23
Printing the Results of a Library Search.................................................................................... 11-23
Copying to and from the Windows Clipboard........................................................................... 11-24
To Copy a Picture to the Clipboard................................................................................ 11-24
To Copy the Current Hit List to the Clipboard............................................................... 11-24
To Retrieve Data from the Clipboard............................................................................. 11-24
Refining the Search Spectrum ................................................................................................... 11-24
General ........................................................................................................................... 11-24
To Refine the Search Spectrum...................................................................................... 11-25
Auto Refine .................................................................................................................... 11-25
Library Compare Process .......................................................................................................... 11-25
General ........................................................................................................................... 11-25

Page xiii
MassLynx NT Users Guide

To Compare the Search Spectrum with a Particular Library Entry................................ 11-25

Library Subtract Process ........................................................................................................... 11-26
General........................................................................................................................... 11-26
To Subtract a Particular Hit from the Search Spectrum ................................................. 11-26
User Libraries ............................................................................................................................ 11-26
General........................................................................................................................... 11-26
To Create a New User Library ....................................................................................... 11-26
To Append a Spectrum to a User Library ...................................................................... 11-28
To Add Spectra from a Library to an Existing User Library.......................................... 11-28
To Create a Spectrum and Add it to an Existing User Library....................................... 11-29
Adding Textual Data to Library Entries......................................................................... 11-29
Indexing a User Library ................................................................................................. 11-31
Deleting Library Entries................................................................................................. 11-32
The Library Locator .................................................................................................................. 11-32
General........................................................................................................................... 11-32
To Select a Particular Entry for Display ........................................................................ 11-33
To Locate Entries with Filters........................................................................................ 11-33
To Set the Locate Filters ................................................................................................ 11-33

Chapter 12 Molecular Mass Calculator......................................................................................12-1

Overview ..................................................................................................................................... 12-3
To Calculate the Molecular Mass for a Given Chemical Formula .............................................. 12-3
Defining User Elements ................................................................................................... 12-4

Chapter 13 DataBridge ................................................................................................................13-1

Introduction ................................................................................................................................. 13-3
To Convert a File with DataBridge ............................................................................................. 13-3
General............................................................................................................................. 13-3
Starting the DataBridge Program from the Windows Start Menu.................................... 13-3
Running the DataBridge Program from the MassLynx Sample List................................ 13-6
To Convert an ASCII File to MassLynx Format.............................................................. 13-7

Chapter 14 AutoLynx...................................................................................................................14-1
Introduction ................................................................................................................................. 14-3
Starting AutoLynx....................................................................................................................... 14-3
AutoLynx Settings....................................................................................................................... 14-4
General............................................................................................................................. 14-4
AutoLynx Settings Dialog: Directories Page ................................................................... 14-4
AutoLynx Settings Dialog: Control Page......................................................................... 14-5
AutoLynx Settings Dialog: Operations page.................................................................... 14-6
AutoLynx Settings Dialog: Results Page ......................................................................... 14-6
Interfacing with External Programs............................................................................................. 14-7
Batch Queuing.................................................................................................................. 14-7
Batch Completion ............................................................................................................ 14-7
Monitoring the Queue Status....................................................................................................... 14-7
Aborting the Queue ..................................................................................................................... 14-7
Accessing Results........................................................................................................................ 14-8
Directory Usage........................................................................................................................... 14-8
File Usage.................................................................................................................................... 14-9
File Structures ............................................................................................................................. 14-9

Chapter 15 Accurate Mass Measure ..........................................................................................15-1

Introduction ................................................................................................................................. 15-3
Manipulating Files....................................................................................................................... 15-4
To Highlight Files ............................................................................................................ 15-4
To Select Files.................................................................................................................. 15-4
To Deselect Files.............................................................................................................. 15-4
To Change the Output Filename ...................................................................................... 15-4
To Change the Mass Measure Parameters ....................................................................... 15-5
To Change the Mass Filter Parameters............................................................................. 15-5

Page xiv
 MassLynx NT Users Guide

To Perform Secondary Reference Correction .................................................................. 15-6

To Change the Current Directory..................................................................................... 15-7
Process Type .................................................................................................................... 15-7
Processing Files ........................................................................................................................... 15-7
Closing Accurate Mass Measure ................................................................................................. 15-8

Chapter 16 Installation Qualification Checker.......................................................................... 16-1

Introduction ................................................................................................................................. 16-3
Installation Checking................................................................................................................... 16-3
Accessing the IQ Checker; the Installation Qualification Checker Window............................... 16-4
The Installation Qualification Checker Menu Bar....................................................................... 16-4
The File Menu .................................................................................................................. 16-4
The View Menu................................................................................................................ 16-5
The IQChecker Menu....................................................................................................... 16-5
The Help Menu ................................................................................................................ 16-5
The Installation Qualification Checker Tool Bar ........................................................................ 16-6
Getting Started............................................................................................................................. 16-6
To Open an Existing IQ Check File ................................................................................. 16-6
To Save an IQ Check file ................................................................................................. 16-6
To Change the Font of an IQ Check Report..................................................................... 16-6
IQ Checking ................................................................................................................................ 16-7

Page xv
MassLynx NT Users Guide

Page xvi
 Introduction

Chapter 1 Introduction

Page 1-1
Introduction

Contents
General .......................................................................................................................................... 1-3
Conventions................................................................................................................................... 1-3
New Features in MassLynx NT 4.0............................................................................................... 1-4
New Features in MassLynx NT 3.5............................................................................................. 1-22
New Features in MassLynx NT 3.4............................................................................................. 1-28
New Features in MassLynx NT 3.3............................................................................................. 1-31
New Features in MassLynx NT 3.2............................................................................................. 1-33
New Features in MassLynx NT 3.1............................................................................................. 1-35
New Features in MassLynx NT 3.0............................................................................................. 1-36

Page 1-2
 Introduction

General
This User’s Guide is designed to introduce the User to some of the main features of the
MassLynx data system. When the User has read this manual, they should be able to:

• Select and display data files.

• Process chromatogram and spectrum data.

• Quantify data.

• Perform Library searches.

This manual assumes that the User has no previous knowledge of MassLynx, but does have
knowledge of Microsoft Windows.

Conventions
The MassLynx NT User’s Guide follows these typographic conventions:

This: Represents:

bold Menu commands, dialog items, such as push-buttons and text boxes, and
text; e.g. File menu, OK push-button.

italic Place holders for information that must be provided. For example, if asked
to type filename, the User must type the actual name for a file instead of
the word shown in italic type.

Bold italic Keyboard keys; e.g. Esc, Return.

Keyboard Formats

Key combinations and key sequences appear in the following formats:

KEY1+KEY2 A plus sign (+) between key names means to press both keys
simultaneously. For example, “press ALT+ESC” means to press and hold
down the ALT key, press and hold down the ESC key, then release both
keys.

KEY1,KEY2 A comma sign (,) between key names means to press and release the keys
one after the other. For example, “press ALT,F” means to press and release
the ALT key and then press and release the F key.

Page 1-3
Introduction

New Features in MassLynx NT 4.0

This section gives a brief outline of the main enhancements and changes between MassLynx
NT Release 3.5 and MassLynx NT Release 4.0. Each of the features below is discussed in
more detail in the relevant manual.

MassLynx V4.0 will run on Microsoft Windows XP Professional, Windows NT V4 and

Windows 2000; it no longer supports Windows 95 and 98.

Full functionality of all features in MassLynx V4.0 is not available to all instrument
configurations. Where a specific configuration is required, this is indicated in the relevant
section.

Installation

Standardized mechanism for the installation of Windows-based applications, using the

‘Microsoft Windows Installer’.

Runtime Source Resiliency: The installer can dynamically diagnose and repair corrupt or
missing components, without User intervention.

Rollback: In MassLynx NT Release 3.5, installation errors leave the computer in an

intermediate state, future installs may be prevented, and existing products may be broken.
‘Rollback’ provides the ability to return the computer to its fully-working condition prior to a
failed installation.

Flexibility: An existing installation can be modified, in that, new Application Managers can
added without the need to reinstall the entire product.

Uninstall: Allows the computer to be returned to its working condition prior to the installation
taking place.

Installation Wizard: Restructured, simplified, and easier-to-use.

Enhanced Upgrade Mechanism: Streamlined, stable upgrades to the MassLynx product suite.

Inlets

New Setup

A new ‘Inlet Configuration Wizard’ has been added to walk the User through the configuration
of the inlet system. The User is now only presented with the supported configurations for the
pump being used, greatly simplifying system setup.

The following new configurations have been added:

Waters 2487, 2488, SATIN and 996 detectors are supported with Jasco, Shimadzu,
and Agilent pumps.

Shimadzu Pump, Gilson Autosampler, and Agilent 1100 Diode Array Detector.

MUX Pump Control

It is now possible to run four pumps on a 4-way MUX system. Each pump can be setup
individually, and can then be run in parallel from the sample list, each with a different method.

Page 1-4
 Introduction

In addition, it is also possible to collect data from four diode array detectors on a 4-way MUX
system.

This functionality is restricted to the following configurations:

Pumps: 4 x Waters 600, or 4 x Waters 1525.

Autosampler: CTC PAL (with Harney Valve Module, or CTC Multi Valve).

Detectors: 4 x Waters 996, or 4 x Waters 2996, or Waters 2488.

Waters

The Waters CapLC control shipped with MassLynx 4.0 now supports parking of
chromatographic peaks in order to facilitate greatly enhanced DDA analysis. This feature is
available only on the EPCAS Q-Tof instruments, QTOF Micro, Quattro Ultima, and Quattro
Micro.

New Waters devices supported in MassLynx 4.0:

Waters 2747 Autosampler. Waters 2757 Fraction Collector. Waters 2767

Inject/Collect device. Waters 2525 pump (including the Waters CFO). Waters 2488
MUX UV detector.

Note:
This version of software also supports the Waters 2996, when configured as a Waters 996.

CTC

CTC PAL macros are now included, in order to support ‘cherry picking’ or random vial access
on MUX systems.

CTC PAL macros are included to support inject ahead.

The CTC A200SE is now supported.

The CTC GCPAL can be switched into A200SE emulation mode on the device itself. There is
now a MassLynx autosampler selection to support this.

Agilent

New support for Agilent devices:

Agilent Capillary Pump G1376.

Agilent Well Plate Autosampler (including thermostatic option) G1367A.

Agilent Micro Autosampler (including thermostatic option) G1389A.

It is possible to setup custom beds on the Agilent Well Plate Autosampler.

Gilson

New support is included for Quad Z Autosampler. This device can be used with both 849 and
889 injectors on 4- or 8-way MUX systems. The sampler can also be configured to perform
single injections if required.

New support is included for the ‘Nebula’ pumps - the 321 and 322.

Page 1-5
Introduction

FractionLynx

The Gilson 215 is now supported as an Inject/Collect device.

Pot directed fraction collection has been added.

True UV only Fraction collection is now supported.

It is now possible to apply Boolean logic to mixed triggers for collection of fractions.

Shimadzu

New support for controlling the Shimadzu column switching device.

LC Packings

Support control of Ultimate pump and UV system, together with the Famos well plate
autosampler or Ultimate carousel autosampler and the SwitchOS II unit

Instruments

ZQ

Support generation of external signal based on mass trace, in order to drive, for instance, a
fraction collector.

LCT

It is now possible to acquire data from two analog channels per data file on a 4-way MUX LCT

Quadrupoles

The instrument calibration window now has a hardcopy option available, in order to print
details of the currently loaded calibration.

The maximum centroid threshold has been increased to allow the rejection of noise peaks when
performing a centroid acquisition, thus aiding in the acquisition of smaller data files.

Quattro Micro

First full version to support the Quattro Micro.

It is possible to specify a Set Mass in the sample list MASS columns, for use in Daughter or
Parent Scans – thus facilitating ‘walk up’ MSMS.

Q-Tof

First full version to support the EPCAS QTOF 1.

First full version to support the QTOF API, QTOF API - US, QTOF, MALDI, and QTOF
Global.

Support is now included for 4-way Accurate Mass MUX acquisitions on EPCAS instruments
only.

QTOF Micro

First full version to support the QTOF Micro.

Page 1-6
 Introduction

The QTOF Micro software is now compatible with an instrument that is using LockSpray or
MUX.

M@LDI

Support Dual Linear/Reflectron Mode.

Support PAD (high mass detector).

Support robot stacker and bar code reader for use on high throughput systems

GCT

New support for DCI and FI probes

AutoSpec

The AutoSpec NT system now has a new AutoTune facility

The resolution checking facility now includes the ability to print each reference peak examined
on a single page.

New support for DCI Probe Control.

It is possible to control mechanical slits.

Shutdown

A new facility has been added to allow an email to be sent when a system shuts down.

MassLynx General

Main Application User Interface Enhancements

The main enhancements to the main Application User Interface in version 4.0 are:

General Look and Feel

The main application is now intended to mirror the look of the Micromass website. This
includes the ability of almost everything within the main application to display a ToolTip
giving a hint as to its function or its current status.

Rationalized Toolbar and Menu Item Contents

In MassLynx 3.5, the Tool Bar and main application menus could become very cluttered.
Individual Tool Bar buttons were also difficult to identify due to their small size.

The menu and Tool Bar have now been simplified so that only those items pertinent to the
main application are present. The Tool Bar buttons have also been increased in size and are
displayed in full true color.

Live Status Bar

The status bar has been updated to provide quick access to the settings associated with a
particular status item. For example, clicking on the instrument status pane will invoke the
instrument settings dialog.

Page 1-7
Introduction

Sample List Editor

The main User Interface modifications to the Sample List Editor include more rational context
menu contents and the implementation of its own menu bar.

A reference to the current sample list format file (if one is open) is now saved along with the
sample list. This will mean that the next time a given sample list is opened, it will appear with
its associated format.

The autosampler rack layout is now held within a floating window. This will allow the User to
generate/modify a sample list using the control and then dismiss it, rather than keeping it
present next to the sample list

The spectrum, chromatogram and map applications are now accessible by clicking on the
application name, located immediately above the sample list, rather than going through the
menu system, or attempting to identify their given toolbar image.

Shortcut Bar – the New Application Launcher

Applications are now organized into groups (usually denoted by their application manager) and
displayed in the Shortcut Bar. When an application manager is installed on the system, its
group name will appear on the menu displayed to the left of the Shortcut Bar. Clicking on the
item within this menu will cause the Shortcut Bar to be populated with the list of applications
within that group.

Quick Access to Help

When help is available for a particular shortcut group, the appearance of the group banner
within the Shortcut Bar will be changed to include a question mark. If the User clicks on the
banner, they will be sent directly to the help associated with that group.

Revised Queue Status Display

The status of the queue item currently being processed is now always visible in the information
bar at the top of the main application display.

The familiar queue start, stop and pause buttons remain on the toolbar in order to facilitate
quick access to these features.

In order to view or modify the full contents of the queue, the Queue Bar can be displayed. This
will present a graphical representation of the current queue contents. Clicking on an item will
show its current settings and status.

Revised Instrument Status Display

The instrument status display has been modified to include larger graphics and its overall look
has been updated to fit in with the rest of the main application.

MassLynx without MassLynx Security Manager

New BackLynx for scheduling file moving and copying operations.

Instrument and inlet metadata files no longer written to by MassLynx, except by an explicit
save.

MassLynx Security Manager

All known version 3.5 bugs fixed.

Page 1-8
 Introduction

Extra permissions in security manager.

No longer linked to NT screensaver for timeouts.

No longer uses NT Event Viewer for log information.

New LogService for storing audit log.

New LogLynx, the audit trail logging tool, for viewing and maintaining audit log.

Improved QuanLynx audit trail.

MassLynx integrity crash monitor.

Maintenance login mode established for critical interventions.

MassLynx Security Manager with Secure File Access

MassLynx 4.0 provides a number of new security tools. These tools can be used in
conjunction with appropriate SOPs (Standard Operating Procedures) to aid compatibility with
the US FDA 21 CFR 11 regulation

When operating in Secure File Access mode, MassLynx 4.0 monitors the accidental, deliberate
or unauthorized modification of experimental files using encrypted file checksums. These
checksums are unique to each file. The MassLynx security toolset automatically updates these
checksums when any authorized file access is performed. Any modification to a file without a
corresponding update of the checksum will flag the file’s content as insecure.

Full auditing of all MassLynx files, including data files, requires the use of the Micromass
EPCAS (Embedded P.C. Acquisition System) technology.

EPCAS Systems

EPCAS instruments use the real time encrypted checksum (described above) in conjunction
with the MassLynx 4.0 security tools to ensure the integrity of raw data files as they are
acquired.

Non-EPCAS Systems

Non-EPCAS instruments are unable to acquire raw data with real-time checksums. The
prevention of accidental or deliberate modification to raw data, in accordance with 21 CFR 11
requirements, cannot be guaranteed.

Where possible, a migration path to EPCAS will be provided for older systems; this will be
purchasable as an on-site upgrade. For more details, please contact your local Micromass sales
office.

File tampering is made impossible by encrypted checksums immovably attached to files.

New secure files mode, where files are written and read by the MassLynx User, not the desktop
User.

Electronic Record/Electronic Signature functionality fully implemented.

All file access done as the MassLynx User, not the desktop User.

NTFS file ownership reflects last User to write to file (unlike in NT).

Signatures can be forced or applied optionally on data, metadata, and results.

Page 1-9
Introduction

Audit trails on file actions.

Tamper detection on files with contents or location altered outside MassLynx.

In a secure files environment, BackLynx is a requirement to move and copy MassLynx files,
allowing audit information to be kept with archive sets to ensure full file integrity and
validation. The movement of any MassLynx files is disallowed, except by BackLynx. Once
files have been archived by BackLynx from the MassLynx secure files system, they can be
moved by third-party systems such as CD archiving tools. When a file is required for review
in a secure files environment, the MassLynx archive set must first be imported using
BackLynx. BackLynx will at this point validate the file history of all imported files to ensure
data integrity.

Using BackLynx, data can be transferred from one Secure File Access environment PC to
another Secure File Access environment on another PC. An example typically would be for
data to be transferred from a secure instrument PC to a secure remote server, or individual
secure workstation for post-processing.

Legacy data can be imported by project or individual method files. The audit trail of these files
only commences once the data has been imported.

Dual authorization protection for high level actions such as legacy file import.

Ability to disallow network file access.

Chromatogram

Improved representation of pre-cursor ion chromatograms in function switching experiments

Library

A link to NIST MSSearch application has been added.

Elemental Composition

When launched from the MassLynx spectrum window, the active spectrum is copied to
Elemental Composition.

The spectrum displayed in Elemental Composition forms part of the printed report.

Elemental isotope clusters are compared to the spectrum in Elemental Composition and ranked
by goodness of fit.

Interfacing Tools

Datafile Access 2 Component: An ActiveX component for accessing MassLynx data files from
VB or VC++ applications.

BioLynx

PepSeq

New find tag module for manual peptide sequencing.

User-definable BLAST searching parameters.

Page 1-10
 Introduction

CarboTools

Support for User-defined modifications and terminal groups.

MetaboLynx

MetaboLynx Setup and Processing

Property Page Parameter Modifications to Improve Usability

The contents of some tabs have been combined, e.g. Create Spectrum and Spectrum Test are
now together on a tab named Spectrum. The contents of some tabs have been separated, e.g.
Control Sample parameters have been extracted from Unexpected Metabolites, with MS Data
(Unexpected) being moved onto Unexpected Metabolites. Parameters have been renamed in
order to make their meanings more easily understood.

Spectrum Mass Matching for Control Comparison

If a peak is found in both the control sample and the metabolized sample, within the RT and
response window, the mass spectra can now be compared to determine whether the two peaks
are due to the same compound. The region of the mass spectrum that is compared should
depend on the Amu Step size set on the Control Sample page. Parameters have been added to
the Control Sample tab page to define the tolerance in the peak intensities for matching
(Intensity tolerance %) and the peak intensity threshold for peaks to be included for matching
(Intensity threshold %). If all the analyte mass peaks match within the defined thresholds, the
peak should be marked as matched; this will prevent the spectrum from being considered for
unexpected metabolites. If any of the mass peaks do not match, the spectrum is marked as not
matched; the spectrum will now be considered for unexpected metabolites and there will be
some number of entries in the unexpected metabolite list.

Mass Measurement Option to AFAMM Whole Data File or Chromatogram Peaks Only

On the Pre-Process Data tab, in the Mass Measure Continuum MS Data frame, there are
two new radio buttons - Entire data file and Spectra of chromatogram peaks only.

Selection of Spectra of chromatogram peaks only will carry out chromatographic peak
detection on continuum data and then only carry out mass measurement on spectra produced
from detected peaks. If Use accurate mass spectra only is selected on the Spectrum tab,
these rules will be applied.

If Entire data file is selected, the data-file will undergo AFAMM before chromatographic
peak detection takes place. This will speed up data processing if only a small number of mass
chromatograms are to be integrated.

Analog/PDA Trigger for MS/MS

This new option is used to reduce the number of MS/MS functions set up from the MS data
when there are chromatogram peaks from both MS data and Analog or PDA data in the same
data file. If peaks only exist in either MS data or Analog/PDA data, the option is not to be
applied. The new option allows a metabolite list entry to be set up for MS/MS only if a signal
appears in both the MS and the Analog or PDA chromatograms, i.e. if the MS peak has a
co-eluting peak in any of the Analog or PDA chromatograms.

MS/MS Common Product Ions and Constant Neutral Losses Redefined

Redefinition of Common Product Ions: All mass peaks above a User-defined threshold in the
parent spectrum and the currently selected metabolite spectrum should be compared. If any of
these match, within a User-defined mass tolerance, then they should be added to the list. There

Page 1-11
Introduction

will be two values for each match - the product ion mass from the parent spectrum and the
product ion mass from the metabolite spectrum.

Redefinition of Common Neutral Losses: All mass peaks above a User-defined threshold in the
parent spectrum and the currently selected metabolite spectrum should be compared. If any
peak in the metabolite spectrum differs from a mass peak in the parent drug spectrum by the
same mass difference between the precursor ions for the two compounds, within a User-
defined tolerance, then they should be reported. For instance, if the metabolite precursor ion is
80 units mass higher (such as a sulfate conjugation), than the parent drug precursor, then the
ion in the metabolite spectrum which is 80 units mass higher should be reported. There will be
three items of interest - the common neutral loss, the production for the parent drug, and the
product ion for the metabolite.

Speed Improvement in Chromatogram Generation

MetaboLynx has reduced the time taken to generate chromatograms. Where no time range is
specified, the full acquisition time range is used, and generation is twice as fast as Version 3.5.
If a reduced time range is requested, chromatogram generation is proportionally faster, since it
now generates partial chromatograms instead of full range.

Potential Metabolites New Option ‘Excluded Masses’

This new option will allow known contaminants to be excluded without removing nominally
isobaric components of interest. The option Excluded Masses has been added to the Potential
Metabolites parameters. A new dialog box is set up to contain a list of masses (each with an
optional retention time). A User-specified retention time window can be applied to the
Excluded Masses option. A single mass window is be applied to all masses. Any spectrum
mass that matches a mass in the list should not be considered as a potential metabolite - it
should be considered as an Unwanted metabolite, as shown in the spectrum status in the
header. The Unexpected Metabolite view in the Browser now labels these unwanted
metabolites with a question mark.

Support for MS/MS Acquisitions - Data Dependent MS/MS and Collision Profiles Data

MetaboLynx is able to process data from acquisitions using data-dependent MS to MS/MS

switching (automatic function switching). It processes MS/MS data acquired using Collision
Energy Profiles and Precursor Discovery Acquisitions. MetaboLynx recognizes a data file that
has been generated using data dependent MS/MS and processes it as indicated below.

MS Data: The MS (survey scan) function is processed normally as though it were a regular MS
acquisition allowing all of the MetaboLynx functionality to be applied. In the Browser, the
data is displayed in the MS views as normal.

MS/MS Data: MS/MS functions are processed automatically and the data displayed on the
MS/MS views of the MetaboLynx Browser. If the set mass for the MS/MS acquisition
matches a metabolite mass found in the MS data at the same retention time, the MS/MS entry
should have the same designation (metabolite name, etc.) as the entry in the MS metabolite list.
If the MS/MS set mass does not match with any peak found in the MS data, it is labeled
appropriately in the spectrum header.

Fix for EleComp Results Different from Spectrum EleComp

Sometimes Elemental Composition gave different results from Spectrum, when lengthy
compositions were being done.

Page 1-12
 Introduction

MetaboLynx Browser

Fix Saving of Report Header Text Information in Parameter File on Browser Exit

When formatting a MetaboLynx Report Scheme (*.mrs), any information entered into the
Report Header Text field on the Report Header page of the Edit Report Scheme Settings
dialog was lost as soon as the MetaboLynx Browser was closed. This occurred even if the
*.mrs and *.rpt files were saved first.

Report Metabolite Results to Electronic File

It is now possible to generate a User-defined .TXT file containing all the contents of the
Metabolite Table report. The command is available as File, Save Metabolite Table.

Elemental Composition and MS/MS Fields Removal by Right-Mouse Click

It is now possible to remove fields from both the Elemental Composition display and the
MS/MS windows using a right-click on the header bar of the window to access a drop-down
menu. This is the same as already done for the expected and unexpected metabolite lists.

Transfer EleComp PPM and mDa Fields to Unexpected Metabolite View

When elemental composition results are available for entries in the Unexpected Metabolite list,
it is now possible to select an entry in the Elemental Composition list and copy the formula of
the elemental composition to the relevant field in the Unexpected Metabolites list. This is
achieved by using a right-click on the elemental composition entry, then choosing the relevant
command from the drop-down menu.

Update PPM and mDa Fields on Change of Formula

If the elemental composition of an entry in the unexpected metabolite list is modified, the PPM
and mDa error columns are recalculated by comparing the calculated mass from the Elemental
Composition with the mass measured from the mass spectrum.

Metabolite Views - %Area Calculations on the Fly

The % Area fields of both Expected and Unexpected Metabolite views (MS, analog and PDA
where applicable) are now recalculated and updated whenever a metabolite’s Status field is
changed. If an entry in the list is changed from Found to either Unexpected, or Not found, it
is no longer included in the calculation of % Area.

Open Last Processed Report File

The Browser can be launched with either the last processed report file, the last read file, or
neither. The options are on the File menu.

Combined Detected Metabolite Chromatogram

This new chromatogram can be displayed at the top of the chromatogram window; it is
displayed above any Analog 1 or PDA TAC channel. The chromatogram is a combination of
all of the chromatograms that contain detected peaks appearing in the expected and unexpected
metabolite lists. There are options to Include expected metabolites only, Include all found
metabolites and Include all detected peaks.

Include expected metabolites only will display a combined chromatogram with all of the
detected peaks in the expected metabolite list that have been designated as Found.

Page 1-13
Introduction

Include all found metabolites will display a combined chromatogram with all of the Found
peaks form the expected metabolites list and all of the entries in the unexpected list that have
been edited to be designated as Found.

Include all detected peaks will display a combined chromatogram with all of the Found peaks
form the expected metabolites list and all of the entries in the unexpected list that are
designated either Found or Unexpected.

Autostart MS/MS option to ignore MS/MS masses if too many

When running AutoStart MS to run both MS and MS/MS sample lists, the browser can now be
set up to not display the dialog box asking for confirmation when there are too many masses.

Molecule Viewer

Sample List Structure Field Supported by MetaboLynx

It is now possible to enter the structure of the parent drug in the Structure column of the
sample list. The structure is entered as the name of a .mol file. Presently only the parent drug
structure can be entered.

Structure Window in MetaboLynx for Parent Drug

A structure window has been added to the browser. This window displays the structure of the
currently selected metabolite, if available. Presently, only the parent drug structure can be
displayed.

MetaboLynx Browser Structures Printing

Available structures can be printed with the relevant metabolite. The MetaboLynx report
scheme settings now has a structure control tab with the option to turn structure reporting on or
off and to specify the size and location of the structure in the report.

OpenLynx

Setup

Simplify Method Setup Pages

OpenLynx Setup can now be configured to suit User requirements. Irrelevant tab pages can be
removed to leave a targeted OpenLynx Setup configuration. A configuration wizard is invoked
on initial Setup access after OpenLynx installation. This provides a brief description of what
each tab page is used for, and gives the User the option to hide unused pages. Subsequently,
these pages can be displayed or hidden via the View, Options menu.

OpenLynx Login

Faster Single Page Walk-up

OpenLynx Login now provides two login modes; the existing wizard mode and a new single
dialog mode that collects all the requisite login information. This single dialog is similar to the
original ‘Single Page’ login concept.

Order OpenLynx Methods in OA Login Wizard

The list of OpenLynx methods displayed during login can be ordered alphabetically or by date.
The OA Login administrator may modify this setting.

Page 1-14
 Introduction

Browser

Date and Time Displayed in Browser

The date and time of sample acquisition is now displayed in OpenLynx Browser. This is the
sample acquired date and time information that has been stored in the OpenLynx report file.

OpenLynx Global Server Version 1.0 (OLGS)

New product released with MassLynx V4.0.

OpenLynx

Processed Data Database Storage

Enables OpenLynx processed sample data to be inserted directly (via OLGS) into a pre-defined
set of Oracle database tables. Insertion is done on a per sample basis, NOT per report file. The
system supports multiple concurrent sample inserts from more than one client OpenLynx
workstation.

Flexible Web Server Directed Database Access

More than one OLGS database can be created, and sample data from specific lab machines can
be directed to nominated databases through the OLGS web server by changing settings in the
OpenLynx Setup page.

Sample Status Monitoring

Optional status messages can be written to an OLGS sample insert Log file. Through the
OpenLynx Setup tab for OLGS, the User can enable the email facility for OLGS sample insert
failure and direct the email to a system administrator or similar. The User has full control over
what action to take when a duplicate sample batch ID/sample ID pair is found in the database
prior to insertion. Duplicate samples may be not inserted, used to overwrite all duplicates, or
appended to the database as a new sample. This is important when re-processing data with the
OLGS option enabled.

Note:
Report file generation functionality is not affected.

OpenLynx Global Server

Customizable User Settings

OLGS Login page for individual User accounts per client machine. The User profile page
enables changing of User account details such as password. Cookie storage technology is used
to save and restore individual User customized parameters according to login information.
Navigation bar allows easy access to all settings/parameter pages at any time from the main
User Interface.

Web Based OLGS Search/Data Mining Tools

Common Search Page

Presents Users with a quick and easy way of retrieving sample reference information from the
OLGS database. Fixed table/column configuration is used to search the database on generic
columns such as Sample ID, Batch ID, Submitter, Chemist name.

Page 1-15
Introduction

Advanced Search Tool

Provides an easy means for fine grained data mining. Page displays drop down combo boxes
that let the User choose which database Table, Column, Operator and Value search criteria to
use. These can be combined with Boolean (AND/OR) logic to generate very complex queries
that can be used to narrow the search results as desired.

Search Results Summary

The Sample Search Results page displays a summary of generic sample information of the
samples found matching the current search criteria. Results are paginated to display a
manageable number of samples on one screen. An indication of the total number of samples
returned by the query is also displayed along with a date/time stamp of when the query was
executed. This summary page is printable.

Web Based OLGS Results Browser Components

General

The familiar look and feel of Diversity Results Browser has been maintained but all
components are Java based. Java Applets are used to provide interactive graphical components
such as Microtitre plate, Spectrum and Chromatogram plots.

Interactive Microtitre Plate Applet

Displays all samples in the current sample’s batch in the plate configuration specified (if any).
Multiple plates are supported. The User can change well selection via the mouse and/or cursor
keys.

Tabulated Results Panels

All results tables are customizable in terms of which columns are displayed and what decimal
point format is used for numeric values. These settings are controlled via individual setup
pages accessed through the OLGS Navigation bar.

OLGS Results Panels include:

1. Sample Reference information.

2. Sample Fraction Collection information.

3. Interactive Spectral Peak List pane for current sample.

4. Elemental Composition information pane for current peak.

5. Spectrum Library Search information for current peak.

6. Spectrum Applet including:

a. Full ‘rubber banding’ zoom capability.

b. Customizable peak and axes annotation.

c. Number of decimal places formatting.

7. Chromatogram Applet including:

a. Full ‘rubber banding’ zoom capability.

Page 1-16
 Introduction

b. Customizable peak and axes annotation.

c. Baseline drawing for peak integration.

d. Optional Peak shading for peak integration.

e. Number of decimal places formatting.

OLGS Database Features

Automatic Database Creation

Database schema is created and initialized at OLGS installation using a series of SQL script
files. Enterprise Java Bean (EJB) technology is used within the database itself for seamless
connectivity and data integrity. Automatic EJB deployment takes place during OLGS
installation.

Flexible, but automatic, creation of effective OLGS table space layout also takes place at
installation. The software will suggest a file location for the database files, but this can be
modified by the User at installation.

OLGS generates a unique internal reference system for samples and initiates extensive table
indexing for query optimization. Automated scripts for OLGS database table maintenance are
also supplied as part of the standard installation.

ProteinLynx

PeptideAuto

Support for ProteinLynx Global Server 1.1.

Support for search results archiving in the Bio-Rad WorksBase system.

ProteinProbe

Support for ProteinLynx Global Server 1.1.

Support for sequence tag and composition searches.

Support for search results archiving in the Bio-Rad WorksBase system.

ProteinLynx Global Server V1.1

Search Engine

Unix support.

Support for Tag/Composition and Sequence searches.

Enhanced performance.

Improved scoring.

Improved error support.

Page 1-17
Introduction

Databank Management Tools

Improved administrator tools.

Bio-Rad WorksBase Support

Support for results archiving in the Bio-Rad WorksBase system.

Bio-Rad sample plate tracking.

QuanLynx

General

Electronic Records and Signatures are supported when the Secure File Access mode of
Security has been installed on an EPCAS instrument.

Quantify Method and Calibration fields available Sample List. If entered, these fields take
precedence over files specified in the Create Dataset dialog. Allows multiple methods to be
used within a single batch, samples are grouped by Method.

PK Information fields “Subject Text” and “Subject Time” added to Sample List, fields are
available for reporting in QuanLynx summary.

COM interface implemented to support secure output of LIMS information to third party
applications.

Command line interface added to enable automated reporting and LIMS output of QuanLynx
Dataset information.

QuanLynx automation supported by AutoLynx.

User Interface

“Wait” cursor displayed when QuanLynx is busy.

Summary table numeric information formatted by Decimal Places, Significant Figures, or

Scientific Notation.

Peak Annotation can include concentration and primary/secondary peak ratio information.

Calibration name and timestamp appear in Calibration Window title.

Current Sample ID and Text displayed in Information Bar when displaying summary by
sample.

Peak Modifications automatically saved when selecting new chromatogram.

Current chromatogram display range maintained when modification saved.

User can manually Flag a chromatogram peak.

Summary Table selection maintained when switching between Groups.

Display/editing/reporting of multiple peaks on a chromatogram.

Extra output columns made available in Totals Bar and Totals report.

Page 1-18
 Introduction

Processing

QuanLynx processing supports multi-injection data files.

Support for Multiple Internal Standards.

Calibration Curve Fit algorithm used by Waters Millennium system adopted.

Estimated Maximum Possible Concentration supported.

Toxic Equivalence Factors supported.

Secondary ion peak flags can be displayed in summary.

Automatic retention time update takes current RT order into account.

Totals Groups can be split over multiple acquisition functions.

Calculations added to convert measured Isotope Ratios into mole ratios.

Reporting

Sample report format allows selection of chromatograms, print order and RT ranges. Position
of page breaks can be specified to allow User-defined pagination.

Compound Summary Report allows compound calibration to be reported, enabling compound

summary and calibration to appear on a single page.

Option to disable printing of summary table in Sample Report.

Enhanced User selection of Compounds, Samples, and Groups to be included in report.

User selection of Data file sample information for inclusion in Report sample headers.

Dataset Audit Trail printed report.

Number of Peaks contributing to Totals entry can be displayed in Summary.

QuanOptimize

Quantitation

Sample List Columns

The quantitation method and curve columns are now visible from the sample list. QuanLynx
uses these columns for processing if they are populated, which means that multiple quantitation
methods can be specified for a particular sample list.

QuanOptimize; Optimization Stage

Method Creation

If optimization is performed separately from the analysis stage, an acquisition and quantitation
method is still created for each group.

The method filenames are based on the name of the compound sample list and the group label
involved. For quantitation methods, QM_ is added to the front of the filename, and for
acquisition methods, MS_ or MSMS_ is added, depending on the type of method involved.

Page 1-19
Introduction

Example filenames include: QM_ CompoundList _A.exp, MSMS_CompoundList_A.mdb.

Optimization without Analysis

Compound optimization can now be run without having to specify an analytical sample list. In
this situation, analytical methods are still created, based on the compound groups involved.

Access to Optimization Chromatogram Data

As for a normal sample list, the chromatogram for each compound can be displayed by
selecting the compound in the sample list and then the chromatogram toolbar button.

Optimization of all Compound Groups before the Analysis Stage

There are now two options for running ‘optimization with analysis’:

1. It can be run on a per group basis, as done before.

2. There is now the possibility of optimizing all groups before running any part of the
analysis stage.

These options are set within the QuanOptimize method on the first tab page.

Optimization Restart Option

QuanOptimize now has a restart option that can be found on the first page of the acquisition
wizard. This option enables the optimization process to be restarted, without repeating data
acquisition.

Optimize New Compounds Only Option

There is a now the option to optimize only the new compounds that appear in the compound
sample list.

Optimization Results File

The optimization results file is no longer temporary, but is saved to the current project, with a
filename based on the current compound sample list.

QuanOptimize; Analysis Stage

Method Creation

The quantitation and acquisition methods created for the analytical sample list are now saved
permanently, and the filenames are referenced in the sample list.

All the analytical methods are created at the beginning of the analysis stage, so that, if
necessary, the sample list can be restarted, by running it as a normal quantitation sample list,
i.e. without having to repeat QuanOptimize.

These methods can be viewed and modified as desired, so that the analytical list can also be
used as a starting point for the creation of other quantitation experiments.

The filenames of the methods created are based on the name of the sample list, and the
compound groups involved. If there are multiple groups specified for a particular sample, the
group names are concatenated into the filename, to make it clear which groups make up each
method.

Page 1-20
 Introduction

The acquisition methods will have MS_ or MSMS_ prefixed to the filename, to distinguish
them as acquisition files, and to clarify their type. The quantitation methods have QM_
prefixed to the filename, to distinguish them as quantitation methods.

The QuanOptimize results file is also based on the name of the sample list. If the results file
already exists then an incremented number is added to the end of the filename to prevent
previous results from being overwritten. The acquisition methods and quantitation methods
also have the unique number added to their filename.

Examples of the filenames involved, for a sample containing groups ‘A’ and ‘B’, are as
follows: MSMS_AnalyticalList_AB_001.exp, QM_AnalyticalList_AB_001.mdb.

Analysis Stage Method Creation without Data Acquisition

It is now possible to generate the acquisition and quantitation methods for the analysis sample
list, without starting the data acquisition.

QuanOptimize Method File References

When a new project is created from a template, all the methods used in QuanOptimize are also
copied to the project.

The file references in the QuanOptimize method are updated to point to this new project.

It is still possible to reference files in a different project using the option ‘Reference Files
Outside current project’, from the QuanOptimize Run Wizard.

QuanOptimize Wizard Changes

The QuanOptimize acquisition wizard has been changed considerably to reflect the new
options available, and some of the original options have been removed that where no longer
relevant to QuanOptimize.

The wizard has been reduced to two pages.

QuanOptimize Method Editor

The only new feature in the QuanOptimize Method Editor is the option to perform
optimization on a ‘per group’ or ‘all before analysis’ basis. This is set on the Optimization tab
page.

Open Access QuanOptimize

OpenLynx Methods

QuanOptimize experiments can now be submitted through Open Access Login. To enable this,
an extra page for QuanOptimize has been added to the OpenLynx method editor.

The OpenLynx method is used to specify one particular set of parameters for a QuanOptimize
experiment. An OpenLynx method should be created for each different type of QuanOptimize
experiment required by Users. For example, one method could be used to run compound
optimizations only, while another could be used to perform optimization and analysis. In each
case, the QuanLynx method specified should be suitable for the compounds and samples
involved.

Sample Batch Login

Open Access Login handles submission of QuanOptimize experiments in the same way as
normal sample batches, except that both a compound and analysis sample list must be supplied.

Page 1-21
Introduction

This can be done either through a tab delimited text file, or by simple cut and paste from a
suitable file (simple text, Excel, MassLynx sample list, etc.).

A current restriction is that Open Access QuanOptimize is only configured for whole plate
login, but single sample login will be developed in the future.

A User email address can be specified where the Quantitation results can be sent. For
OAQuanOptimise, a normal Quantation *.qld file is generated, not a normal OpenLynx *.rtp
file.

A directory can also be supplied, where a copy of the Quantitation results will be saved. This
includes the compound optimization results as well as the quantitation report.

Open Access Quantitation

Open Access Quantitation is a method to run Quantitation analyses through the open access
login system.

The conditions required for a particular quantitation analysis are stored within a specific
OpenLynx method, which is selected by the User during the login process.

The OpenLynx method controls the acquisition parameters (LC method, tune file, etc.), the
quantitation options (integrate, calibrate, quantify, etc.) and the quantitation method. The User
can therefore select a particular set of experimental conditions, by simply choosing the
appropriate method; only the sample list has to be supplied.

New Features in MassLynx NT 3.5

This section gives a brief outline of the main enhancements and changes between MassLynx
NT Release 3.4 and MassLynx NT Release 3.5. Each of the features below is discussed in
more detail in the relevant manual.

Supported Operating Systems

MassLynx V3.5 is supported on:

• Windows 2000 service pack 1.

• Windows 98 second version (stand-alone version for data re-processing only).

• Windows NT service pack 6.

LCT

Support for Lock Spray has been added.

Support for Exact Mass MUX has been added.

Support for Pos/Neg switching on MUX has been added.

QTof

Flow can be stopped when switching from MS to MSMS.

Page 1-22
 Introduction

Quattro Ultima (EPCAS)

Enhanced function switching, with include, exclude and adduct lists, collision energy profile,
multi precursor switching and charge state recognition.

Divert valve control in solvent delay.

ZQ

Support for new single quadrupole mass spectrometer.

AutoSpec NT

Autotune now implemented.

Analog channel acquisitions are supported.

FD/FI mode supported.

Full solids probe control.

GCT

Full solids probe control

Waters

Waters CapLC — Pump stop flow for Q-Tof MS-MS/MS switching.

Waters 1525 and 515 — New solvent delivery systems.

Waters 2690, 2790, 996 PDA — New functionality & Control.

Waters 2487 — Digital data collection via IEEE.

Gilson

Gilson 215 — improved control/synchronization with PDA.

Gilson 215 inject ahead.

Shutdown

Functionality has been extended to enable the User to shut down a LC Pumping System in the
case of mass spectrometer errors, such as loss of communications, or lack of gas flow. It is
also now possible to run different context dependent shutdown routines.

HP6890

PTV injector control has been added.

Full dual inlet support has been added.

Page 1-23
Introduction

Jasco

LCNET II support has been added.

CTC PAL

Support for Harney Valve Module – 4-valve injection system.

MassLynx General

A new MassLynx Security Manager, incorporating the Microsoft Windows NT security model,
has been added.

Alternative mode of chromatogram peak integration: ApexTracking.

Optional automatic determination of chromatogram noise for Standard Peak integration.

MassLynx has an automatic link to the Advanced Chemistry Development's (ACD labs)
SpecManager software suite for structural elucidation.

DataBridge

DataBridge can now be executed from the MassLynx sample list thereby enabling batch
processing of data files.

Extra conversion features for AutoSpec Opus - MassLynx with respect to file management
layout.

Quantify

Automatic Limits of detection calculation.

Improvements for Dioxin quantification have been added, including improved reporting of
statistics.

Record Quan calibration parameters in the report.

Page numbering fix for incorrect multi-page sample list results.

Report format Save and Restore as named files.

Sample Information now available in Summary Report when compound peak has not been
identified.

OpenLynx

Standalone HPLC, this option is only supported with detectors which can produces the
MassLynx raw data files, i.e. the Waters 996 PDA and Waters 2487 with the IEEE interface for
collecting data.

Automatic monitoring of system performance.

Compound confirmation based on chromatogram peak purity.

Page 1-24
 Introduction

MaxEnt 1 data processing can be automated with OpenLynx.

The MassLynx all file accurate mass measurement processing can be automated from
OpenLynx.

The MassLynx isotopic cluster analysis processing can now be automated from OpenLynx.

Separate thresholding for compound Spectrum Test.

Up to 30 compounds can now be targeted using OpenLynx.

All sample logging now has an option to submit the samples by file rather than typing the
information via the keyboard.

OpenLynx Login sample status.

The OpenLynx Browser views to may now be copied to the Windows clipboard.

MetaboLynx

The automatic starting of MS/MS Auto-start MS/MS Acquisition.

MS/MS data correlation.

Improved Electronic reporting.

Isotopic Cluster Analysis.

Accurate Mass Spectra implementation as OpenLynx.

Mass Measurement as option on Pre-process page option.

Elemental Composition Analysis for Unexpected Metabolites.

PPM and mDa Calculations for each Metabolite with a Formula.

Eliminate Isotope Entries from Unexpected Metabolites.

Support EPCAS MS/MS Acquisition Method Generation.

Browser Metabolite View Changes:

MS/MS Metabolite Field to be Changeable.

Unexpected Metabolite Name and Formula to be Changeable.

MS/MS Correlation Parent to be changeable.

Status Values in Unexpected Metabolite View.

Decimal Places for Mass Difference and m/z Found.

Mass Difference Calculation.

View Options Metabolite View Hide Not-Found Metabolites.

Redundant Columns Found, Expected.

Page 1-25
Introduction

Select Order of Columns.

Browser Display Range on Control Window to set Analyte Window.

DAD and Analog Peaks in Browser.

Halogens not allowed if none in Parent.

FractionLynx

Chromatogram indication of location of detected fraction regions.

Support Gilson 215 Fraction Collector.

Support Gilson 204 Fraction Collector.

No Mass Spec fraction collection.

Mixed detection modes.

Fraction Collection Location Specification.

Stop injection when fraction collector beds full.

MicrobeLynx

New application manager to enable the rapid screening of Microbiological samples in

conjunction the new M@LDI mass spectrometer.

ProteinLynx Global Server 1.0

Major new product released along with MassLynx 3.5.

Features scalable high-speed database search engine running on multiprocessor systems

enabling true high throughput proteomics.

Databases searched “on the fly” - index files not required.

Improved probability based scoring for both MALDI and MS/MS searches.

Three tier client server architecture, using the Internet and HTTP.

Supports both traditional GUI and Web browser based clients.

Support for both protein and EST databases in FASTA format.

Fixed and variable modifications.

Molecular weight and pI restrictions.

Digest and secondary digest rules with partial digests.

Web browser based server administration.

Page 1-26
 Introduction

Web browser based database searching

Major feature of ProteinLynx Global Server 1.0.

Set-up and submit query via a Web browser to ProteinLynx Global Server.

Results displayed either in terms of matched proteins, matched peptides, or a 'best hit' view,
displaying the smallest set of proteins required to account for all of the submitted query
masses.

Collapsible hit lists.

Spectrum browser tool for displaying and manipulating submitted spectra and deconvoluted
query spectra. Provides graphical representation of the protein or peptide match.

BioLynx

MaxEnt 3 — Charge state restriction below a certain mass.

CarboTools — new application for the interpretation of carbohydrate data:

User configurable for different monosaccharide residues.

Automatic assignment of monosaccharide compositions to loaded spectrum peaks.

Manual assignment of monosaccharide sequences.

Support for derivatives and modifications.

Support for Sodium and Potassium adducts.

ProteinLynx

Client application to ProteinLynx Global Server 1.0.

Greatly increased speed in database searching.

True high throughput proteomics tool.

Simplified single page query set-up.

Query set-up common between ProteinLynx set-up and ProteinProbe.

Support for exclude masses, including autolysis, matrix and lock mass peaks or other masses
defined in a text file.

Support for survey scan processing — automatic generation and saving of integrated peaks.

ProteinProbe

BioLynx Database Searching and ProteinLynx Results Browser. Client application to

ProteinLynx Global Server 1.0.

Greatly increased speed in database searching.

Page 1-27
Introduction

Extremely easy set-up — Set a URL to that of ProteinLynx Global Server.

Simplified single page query set-up.

Query set-up common between ProteinProbe and ProteinLynx set-up.

Entire MALDI spectrum submitted, no User peak picking required.

Single probability based score value.

Support for exclude masses, including autolysis, matrix and lock mass peaks or other masses
defined in a text file.

Sequence tag, sequence composition and text searches not supported.

New Features in MassLynx NT 3.4

This section gives a brief outline of the main enhancements and changes between MassLynx
NT Release 3.3 and MassLynx NT Release 3.4. Each of the features below is discussed in
more detail in the relevant manual.

MassLynx

Samples can be added to the sample list using the generic sample list plate loader displayed on
the top level MassLynx screen. This option is available for Waters 2690, Waters 2790 and
Gilson systems.

The Micromass Web site can now be accessed from the MassLynx top-level screen.

Enhanced VB interfacing guide with improved examples of how to access MassLynx data.

BioLynx

ProteinProbe interface now has interactive client server database searching capabilities.

MassSeq – a new MS/MS de novo sequencing program.

ProteinLynx

Automated MaxEnt 3 processing of MALDI and ElectroSpray data is now available.

ProteinLynx results Browser and protein database search engine are now fully integrated
allowing searching and re-searching of unmatched masses.

Automated MS/MS database searching added, with the facility to view results in the browser.

ProteinLynx can now generate BioRad format files for ProteomeWorks integration.

OpenLynx

Improved automatic accurate mass calculation software for the LCT.

Elemental composition parameters file can be loaded into the OpenLynx Method program and
a restricted search can be performed.

Page 1-28
 Introduction

Improved reporting capacities including accurate mass error reporting and the option to print in
Portrait or Landscape format.

NeoLynx

Redesigned Test File Editor.

NeoLynx Browser added for viewing and printing results.

MetaboLynx

Option to target unexpected metabolites added.

Comparison data can now be displayed on the Chromatogram.

Support for Q-Tof Instrument MSMS development based on the results of initial Metabolite
identification.

FractionLynx

Control of the Waters Fraction Collector II added.

All File Accurate Mass Measure

This utility has been extended to allow Secondary Reference Correction and Peak Filtering.

Combine All Files

Combine All Files is used to combine a group of files that have been acquired using the same
acquisition method to produce a single output file. The combination of the data in this way
results in an increase in the signal to noise ratio.

Calibration

Option of lock mass correction on calibrations.

New instrument calibration file format.

Waters CapLC

Control software for the Waters CapLC Autosampler and PDA detector added.

Waters 2700

Waters 2700 spotting device control added.

Jasco Systems

Control software for the Jasco 1500 HPLC Autosampler and UV detector added.

Page 1-29
Introduction

CTC PAL Autosampler

Improved CTC PAL autosampler control. Including method generation using the Cycle
Composer.

MassLynx CTC PAL is now available on the LCT and Q-Tof instruments. OpenLynx also
supports this autosampler.

Support Removed

MassLynx V3.4 does not support the MicroTech LC systems.

The combination of the HP5890 GC and HP6890 autosampler is not supported by MassLynx
V3.4. The HP6890 GC is still supported for use with the HP6890 autosampler, and the
HP5890 GC is still supported for use with the CTC A200S and HP7673 autosamplers.

MassLynx V3.4 does not support the CTC A200S LC autosampler.

The existing MassLynx V3.3 support of the CTC PAL, which used the A200S LC emulation,
is not supported by MassLynx V3.4. It has been replaced with the Cycle Composer
implementation.

Embedded PC Support

MassLynx NT support for non transputer based instruments. This includes a new Tune page
design and operation. (Quattro Ultima, Quattro LC, ZMD, GC-Tof, Maldi, IsoPrime, LCT and
AutoSpec instruments).

MassLynx instrument configuration set-up from inlet editor rather than the instrument control
panel.

MS-MS/MS function switching supported on the Quattro Ultima and Quattro LC.

AutoSpec

Support for ES, APcI, FAB and FI sources added

QTof

QTof now has real-time charge state recognition, real-time peak detection and TDC +ve/-ve
parameter settings

LCT

Control of MUX interface added. Fast acquisition and processing introduced. (At least 10
scans/sec centroid).

MALDI-Tof

Lock mass adjustment introduced.

Page 1-30
 Introduction

IsoPrime

Dual Inlet supported.

Advanced quantification facilities.

A new application for Hydrogen data calculations has been added.

Platform ICP

New periodic table interface for method set-up.

New Features in MassLynx NT 3.3

This section gives a brief outline of the main enhancements and changes between MassLynx
NT Release 3.2 and MassLynx NT Release 3.3. Each of the features below is discussed in
more detail in the relevant manual.

BioLynx

Redesign of the ProteinProbe interface for faster searching.

ProteinLynx

MASCOT compatible output file format added.

AutoSpec Support

MassLynx NT support for the AutoSpec Ultima.

LCT

Sample Cone, Extraction and RF DC One Offset are linked as on other Z Spray machines (use
of old tune files will result in a greater than expected cone voltage being applied).

Vacuum and gas pressures appear on the experimental report.

Readbacks have had ion energy removed.

Steering and Focus readbacks now reflect what goes on in the instrument.

Tune page layout has been simplified. Advanced and Engineers pages can be removed using a
menu option.

It is possible to setup a function specific cone voltage in the function setup.

MetaboLynx

An analytical tool for drug metabolite identification with the ability to view results in a
browser.

Page 1-31
Introduction

QuanLynx

QuanLynx is designed for screening large numbers of samples containing a large number and
variety of compounds. Many different methods are required, with each method often involving
multiple compounds. QuanLynx automatically finds the best cone voltage for transmission of
a parent ion and the optimum collision energy for a given parent daughter transition. These
optimizations are used to create a scan method file used for acquisition and quantitation.

Elemental

An option has been added to the Parameters dialog to display/hide masses whose results do not
fit within the range of calculation parameters.

The parameters for an elemental composition search can now be saved to a file.

Map

Initial mass/wavelengths for the map display are now User definable.

Gilson Pumps

Flow rates can be modified from within the gradient timetable.

Autosamplers

Unique file extensions for bed layouts for Gilson and 2700 autosamplers

Implementation of the Gilson Multi-Injector autosampler. (Gilson 215 autosampler +

Gilson 889 Multiple Injector system).

Strip

There is now an option to process all functions for a data file.

Quantify

When displaying chromatograms in Quantify, the compound primary chromatogram can

appear at the top of the display with secondary chromatogram and IS chromatogram(s) below
it.

Spectrum

Option of color-coded reference peaks in a spectrum.

OpenLynx

Ability to target compounds by relative or absolute amount.

Ability to specify which process to execute from within OpenLynx.

Page 1-32
 Introduction

Option of filename generation in OpenLynx login to specify a rolling filename that is

incremented for each sample submitted.

Errors from the batch processing are reported on the status bar of the OpenLynx Login
program.

The Job ID label text can be changed from the first login wizard page.

HPLC option is now available for Waters 2690 systems.

Option for spectrum thresholding of labels in the OpenLynx Browser.

Option to show or hide the baselines for the results of the chromatogram trace.

OpenLynx Browser has an option to annotate the chromatogram with the Base Peak Mass.

If the browser has no integration information Chromatograms can be annotated with the
retention time.

Improved reporting capacities.

Automatic Instrument Shutdown

An option is now available which enables the User to shutdown the instrument immediately on
an LC error.

Bio-Q/Quattro Family

A new option to automatically generate MRM masses from the masses defined in the sample
list.

Example Macros

Loopproc and Chroproc macros have been changed to allow printing of spectra in landscape or
portrait mode.

Divert Valve support for Platform LCZ/ZMD and Quattro LC

Divert valve can be automatically controlled during acquisition.

New Features in MassLynx NT 3.2

This section gives a brief outline of the main enhancements and changes between MassLynx
NT Release 3.1 and MassLynx NT Release 3.2. Each of the features below is discussed in
more detail later in the manual.

BioLynx

The PepSeqTM de novo interactive MSMS peptide sequencing including find tag.

Page 1-33
Introduction

ProteinLynx

Automated processing of Maldi and ElectroSpray LC/MSMS data for peptide fingerprint
database searching using client server technology.

MaxEnt 3

Massive Inference for deconvoluting MS/MSMS data for peptides and proteins.

OpenLynx

Quantify – Allows parameters to be defined so that OpenLynx can calculate the concentration
and amount of sample based on the areas of peaks detected.

Elemental – Allows elemental composition calculations to be performed.

A new color has been added to the Browser for found tentative compounds.

Accurate mass – allows target analysis of compounds to greater accuracy.

HPLC gradient definition for HP1100.

Support for Waters 2700 Autosampler

MassLynx V3.1 build 6 includes control software for the Waters 2700 Autosampler.

Support for Waters 486 UV Detector

MassLynx V3.1 build 6 includes control software for the Waters 486 UV Detector.

Support for Waters 2487 UV Detector

MassLynx V3.1 build 6 includes control software for the Waters 2487 UV Detector.

Support for Waters 600 Pump

Control software for the Waters 600 Pump series.

Accurate Mass Chromatograms

MassLynx can generate accurate mass chromatograms.

AutoLynx

An application that enables batches to be submitted to the MassLynx queue from a third party
program for acquisition, processing and report generation.

Page 1-34
 Introduction

All File Accurate Mass Measure

This utility allows mass measure and accurate mass measurements (for LCT and QTof data) to
be performed on an entire data file or a selection of data files instead of an individual scan.

Database Logging

Details of all samples acquired can be written to a database to allow machine usage to be
analyzed.

Elemental Composition

MassLynx can produce a list of possible compounds for a given mass or list of masses.

Quantify

A new Export Results to LIMS option to allow the quantification results to be written to a text
file for export to LIMS systems.

Scan Function Editor

A new option to automatically generate SIR masses from the masses defined in the sample list.

Priority Processing

Sample Lists can now be defined as priority processes, which moves the process to the top of
the queue.

Night Time Processing

Sample Lists can now be defined as night time processes to be acquired overnight.

MassLynx Status File

A status.ini file is created that can be viewed across a network allowing Users to decide which
instrument should be used to acquire samples. The file will contain the MS status, the LC
status and details of samples in the queue.

Import Worksheet

Allows sample lists to be written in Access, Excel or Notepad and imported into MassLynx.

New Features in MassLynx NT 3.1

This section gives a brief outline of the main enhancements and changes between MassLynx
NT Release 3.0 and MassLynx NT Release 3.1. Each of the features below is discussed in
more detail later in the manual.

Page 1-35
Introduction

BioLynx

The IndexBuilderTM program builds digest and molecular weight indices for the SWISS-
PROT/TREMBL and any FASTA formatted database such as the OWL database. These
indices allow for faster lookup and are particularly useful for peptide fingerprint searches
where a list of peptide masses (> 5 masses) are used to identify a particular protein.

OpenLynx

Library Searching – Allows Users to define libraries to search results against or to create new
ones from results acquired.

Support for Waters 2690 LC Systems via GPIB interface

MassLynx V3.0 controls the Waters 2690 LC pump via the PC serial interface. MassLynx
V3.1 will control the Waters 2690 LC pump and DAD detector via the GPIB interface.

Support for Waters 996 PDA Detector

MassLynx V3.1 includes control software for the Waters 996 PDA Detector.

New Features in MassLynx NT 3.0

This section gives a brief outline of the main enhancements and changes between MassLynx
NT Release 2.3 and MassLynx NT Release 3.0. Each of the features below is discussed in
more detail later in the manual.

MassLynx top level screen

The “traditional “ MassLynx top level menu bar and Sample List have been replaced by a
single MassLynx top level screen. An instrument status bar has been added to remove the need
to have many windows displayed for routine operation.

Sample Lists

The Sample List is now part of the MassLynx top level screen. It has an MS Access format
and Excel-style cut and paste editing. A new queue management system that allows Sample
Lists can be chained, prioritized, and added during acquisition.

Project Wizard

A project wizard to assist in the creation of projects from templates.

Projects

Acquisition methods are now stored in projects.

Page 1-36
 Introduction

Quantify

The number of calibration standards has been increased to 100. The number of different
concentration levels per sample has been increased to 20.

Longer File Names

Raw data files can now have up to 128 character path/file names.

Analytical Component Engine

A facility to allow the User to change inlet, autosampler and pump without having to re-install
MassLynx.

OpenLynx

Walk up Diversity. Ability to display Chromatograms in the browser. E-mail distribution of

results. Browser for reviewing single sample results. Improved facility for batch run analysis
allowing whole plates to be processed within one simple login session.

Q-Tof

Automated data dependent switching from MS to MS/MS. Accurate mass option.

Chromatogram Peak Purity

An algorithm for calculating chromatogram peak purity.

Chromatogram Signal to Noise

An algorithm for calculating the ratio of the peak heights to the level of noise in a mass
chromatogram.

Chromatogram Retention Index

The Retention Index is used to compare results from different HPLC systems and different
columns.

Support for Gilson Pumps

MassLynx V3.0 includes control software for the Gilson pump.

Support for HP6890 GC Systems

MassLynx V3.0 includes control software for the HP6890 gas chromatograph and autosampler.

Page 1-37
Introduction

Support for Jasco LC Systems

MassLynx V3.0 includes control software for the Jasco LC system including the UV Detector
option.

Support for Waters 2690 LC Systems

MassLynx V3.0 includes control software for the Waters2690 LC system including the UV
Detector option.

Support for CTC/LEAP Technology PAL autosampler

MassLynx V3.0 includes control software for the PAL autosampler.

Support for MicroTech LC Systems

MassLynx V3.0 includes control software for the MicroTech LC system.

Periodic Table

A MassLynx elemental database facility allowing the User access to elemental information for
the whole periodic table of the elements.

BioLynx

A Spectrum has been added to the ProteinProbe window to allow Users to click on peaks to
create queries.

Support for FASTA format databases e.g. OWL.

Ability to search against current results rather than the whole database.

Colors and Fonts

The colors and fonts editor has been changed to allow more colors.

Startup and Shutdown

Automated startup and shutdown procedures.

Macro support

Macro support has been ported to 32-bit Visual Basic and a new 32-bit MassLynx Applications
Program Interface has been introduced.

Page 1-38
 Installing MassLynx

Chapter 2 Installing MassLynx

Page 2-1
Installing MassLynx

Contents
Minimum System Requirements ................................................................................................... 2-3
Installing MassLynx ...................................................................................................................... 2-4
Preparation ......................................................................................................................... 2-4
To Install MassLynx .......................................................................................................... 2-4
Using the MassLynx Installation Wizard ...................................................................................... 2-5
General............................................................................................................................... 2-5
Tool Tips ............................................................................................................................ 2-5
The MassLynx Security Options and Backup Scheduler Dialogs...................................... 2-5
The MassLynx Options, MassLynx Installation directory dialog ...................................... 2-6
Failed Installation, or Installation Cancelled...................................................................... 2-7
The MassLynx Folder ................................................................................................................... 2-7
Installing Updates to MassLynx .................................................................................................... 2-8
Installing Additional MassLynx Application Managers................................................................ 2-8
General............................................................................................................................... 2-8
Procedure ........................................................................................................................... 2-8
Installing MassLynx Libraries....................................................................................................... 2-9
Installing the SWISS-PROT Database ........................................................................................ 2-10
Uninstalling MassLynx ............................................................................................................... 2-10
Repairing MassLynx ................................................................................................................... 2-11

Illustrations
Figure 2.1 The MassLynx Wizard Welcome Screen.................................................................... 2-4
Figure 2.2 The MassLynx Security Options dialog...................................................................... 2-5
Figure 2.3 The Backup Scheduler dialog ..................................................................................... 2-6
Figure 2.4 The MassLynx Options, MassLynx Installation directory dialog ............................... 2-6
Figure 2.5 The MassLynx Folder ................................................................................................. 2-7
Figure 2.6 The Program Maintenance dialog ............................................................................... 2-9
Figure 2.7 The Uninstall Option: Uninstall Java components dialog ......................................... 2-11
Figure 2.8 Auto repair message.................................................................................................. 2-11

Page 2-2
 Installing MassLynx

Minimum System Requirements

The minimum supported system requirements for running MassLynx V4.0 are:

• Compaq AP200 550 MHz PC with 256 Mb RAM (512 Mb recommended), 9.1 Gb Ultra SCSI
hard disk.

Either:

• English Language Version Windows NT 4.0 with Service Pack 6a.

Or:

• English Language Version Windows 2000 with Service Pack 2.

Or:

• Windows XP Professional.

Note:
Windows XP Home will support the stand-alone version for data reprocessing only.

• Java Runtime 1.3.1.

• Java 3D.

• Internet Explorer 5.5.

Note:
The installation will detect whether the above three components are already installed on the PC; if
any are missing the Installation Wizard will install them before continuing with the MassLynx
installation.

• A VGA (or better) monitor/display adapter supported by Windows NT/2000.

• A Windows compatible mouse.

• A CD-ROM drive.

Page 2-3
Installing MassLynx

Installing MassLynx
Preparation

Note:
It is important that the following procedure is performed before MassLynx is installed.

Before installing a new version of MassLynx, switch the instrument into Standby, remove any
probes and switch off all gasses. It is possible that the instrument may be vented as the new
transputer code is loaded for the first time. The PC must be running Windows NT, Windows 2000
or Windows XP before starting to install MassLynx. Any other programs that are running should
be closed down, including any existing version of MassLynx.

Note:
MassLynx V4.0 may be uninstalled using standard Windows procedures, refer to the “Uninstalling
MassLynx” section, on page 2-10, for further details.

To Install MassLynx

Note:
To install MassLynx, the User must be logged on to an Account that has administrative privileges.

1. Insert the MassLynx for Windows Installation CD disk into the CD-ROM drive; the
installation should start automatically and invoke the MassLynx Wizard Welcome Screen.

Figure 2.1 The MassLynx Wizard Welcome Screen

2. Select the Next > button and follow the on-screen instructions to continue installing the
application. See the “Using the MassLynx Installation Wizard” section for further
information.

The installation program creates a shortcut to the MassLynx folder on the PC’s Desktop, and a
MassLynx folder is placed in the Start Menu, see the “The MassLynx Folder” section, on
page 2-7.

Page 2-4
 Installing MassLynx

Using the MassLynx Installation Wizard

General

The MassLynx Installation Wizard is used to install MassLynx from the MassLynx for Windows
Installation CD, see the “Installing MassLynx” section, on page 2-4. Once the Wizard has been
invoked, the User must follow the on-screen instructions; these are mostly self-explanatory. This
section provides additional information about using the Wizard.

Tool Tips

If the mouse cursor is positioned above an option, or other features in a dialog, a Tool Tip may be
available, providing additional details.

The MassLynx Security Options and Backup Scheduler Dialogs

The MassLynx Security Options dialog gives the option of installing MassLynx Security
Manager and hence, the MassLynx Security System.

Figure 2.2 The MassLynx Security Options dialog

Select the No option if MassLynx Security is not required. Select the Yes option if MassLynx
Security is required; refer to the “MassLynx Security User’s Guide” for details of the MassLynx
Security System and its installation.

If the No option has been selected, selecting the MassLynx Security Options dialog Next >
button invokes the Backup Scheduler dialog.

Page 2-5
Installing MassLynx

Figure 2.3 The Backup Scheduler dialog

If required, select the Backup Scheduler option to install the BackLynx utility; this allows files to
be copied between locations without invalidating any security attached to them, refer to the
“MassLynx Security User’s Guide” for details.

The MassLynx Options, MassLynx Installation directory dialog

The MassLynx Options, MassLynx Installation directory dialog allows the User to select the
disk drive and directory in which MassLynx will be installed; this defaults to C:\MassLynx.

Note:
If an alternative drive is selected, a C:\MassLynx directory will still be created in addition to the
User-selected directory.

Figure 2.4 The MassLynx Options, MassLynx Installation directory dialog

Page 2-6
 Installing MassLynx

Failed Installation, or Installation Cancelled

If the installation fails, or the User cancels the installation before it is complete, the Installation
Wizard will “roll back” the installation, i.e. the PC is restored to its original state.

The MassLynx Folder

The installation program creates a MassLynx Folder in the Start Menu.

Note:
On Windows XP, MassLynx V4 may be accessed from Start, All Programs, MassLynx.

Figure 2.5 The MassLynx Folder

The MassLynx folder contains the following items:

Acquisition User Invokes a Help system, which explains how to acquire data.
Guide

Databridge Invokes the MassLynx file conversion program, see Chapter 13,
“DataBridge” for further information.

IQ Checker Invokes the IQ Checker program, which is used to check the validity of a
MassLynx installation, see Chapter 16, “IQ Checker” for further
information.

Macro User Invokes a Help system, which explains how to use Macros.
Guide

MassLynx User Invokes a Help system, which explains how to use MassLynx.
Guide

MassLynx V4.0 Starts the MassLynx program.

BackLynx Invokes the BackLynx Utility; when operating in a secure MassLynx

environment, this allows files to be moved from one location to another
without invalidating them. See the “MassLynx Security User’s Guide” for
further information.

Note:
BackLynx is only present if selected during MassLynx installation.

Note:
If MassLynx has been installed as an acquiring system on a computer which has a TDAT Interface
card installed, then the computer must be connected to the Mass Spectrometer, or a loop-back
connector must be connected to the TDAT interface board in the computer. Failure to do this may
result in the computer failing to run Microsoft Windows NT.

Page 2-7
Installing MassLynx

Installing Updates to MassLynx

Updates to MassLynx are supplied on a CD or a floppy disk. Update disks contain the latest
program enhancements and fault fixes. Each Update Disk should be accompanied by a set of
instructions that describe the contents of the Update disk and how to install it. Each Update Disk
also contains a file, called readme.txt, which includes the same information.

To install an Update from CD:

1. Insert the Update Disk CD into the CD-ROM drive. The installation should start
automatically and invoke the MassLynx Wizard Welcome Screen.

2. Select the Next > button and follow the on-screen instructions to continue installing the
update.

To install an Update from floppy disk:

1. Insert the Update Floppy Disk into the floppy drive.

2. Select the Windows Start, Run command. The Run dialog is invoked.

3. Type a:setup in the Open: text box.

4. Select the OK button. The MassLynx Wizard Welcome Screen is invoked.

5. Follow the on-screen instructions to continue installing the update.

Installing Additional MassLynx Application

Managers
General

Additional MassLynx Application Managers, not installed during the initial MassLynx
installation, may be installed at a later time, if required.

Note:
If MassLynx was originally installed with the Secure File Access Option dialog, Security File
Access option selected, additional Application Managers cannot be added at a later date;
MassLynx must be uninstalled and then reinstalled with the required Application Managers.

Procedure

1. Insert the MassLynx Installation CD disk into the CD-ROM drive; the installation should start
automatically and invoke the MassLynx Wizard Welcome Screen.

2. Select the Next > button; the Program Maintenance dialog is invoked.

3. Select the Modify option.

4. Select the Next > button; the Application Manager Options dialog is invoked.

5. Select the required options and select the Next > button to complete their installation.

Page 2-8
 Installing MassLynx

Figure 2.6 The Program Maintenance dialog

Note:
In Windows 2000, the procedure may be initiated via the Control Panel, Add/Remove Programs
dialog. Select MassLynx V4 in the list box and select the Change button; a prompt will appear,
asking for the MassLynx Installation CD disk. Insert the disk in the CD-ROM drive and follow the
above procedure.

Installing MassLynx Libraries

Several Libraries are available for use with MassLynx, these include the NIST Library and
Chemical Structures, Wiley Library, Toxicology Library, Carlo Erba Pesticides Library and
Pfleger Maurer Weber Drugs Library.

To install these libraries:

1. Insert the MassLynx Libraries for Windows Installation CD, or floppy disk, into the CD-ROM
or floppy drive.

2. Select the Windows Start button; the Start menu is invoked.

3. Select the Run option; the Run dialog is invoked.

4. Type drive:\setup in the Command Line: text box, where drive is the letter of the CD-ROM
or floppy drive.

5. Select the OK button.

If the NIST Library and Chemical Structures is being installed, a dialog box will appear
offering the Library and Structures options.

6. Select the required options.

7. Select the OK button.

Page 2-9
Installing MassLynx

8. A dialog box is invoked, asking which directory the MassLynx software is installed in. The
default directory is c:\masslynx. The Installation program will copy the files onto the hard
disk. When the installation is complete, a dialog, saying that the system must be shutdown
and restarted for the changes to take effect, is invoked.

9. Select the OK button.

10. Remove the CD, or floppy disk, from the disk drive.

11. Select the Windows Start button; the Start menu is invoked.

12. Select the Shut Down option; the Shut Down Windows dialog is invoked.

13. Select the Restart the computer? option.

14. Select the Yes button.

15. When the PC has restarted, logon to Windows. The installed Library will now be available in
MassLynx.

Installing the SWISS-PROT Database

The SWISS-PROT database is supplied on a CD and is installed in a similar way to MassLynx.

Uninstalling MassLynx
MassLynx V4 may be uninstalled as follows:

1. Insert the MassLynx Installation CD disk into the CD-ROM drive; the installation should start
automatically and invoke the MassLynx Wizard Welcome Screen.

2. Select the Next > button; the Program Maintenance dialog is invoked, see Figure 2.6.

3. Select the Remove option. The Uninstall Options: Uninstall Java components dialog is
invoked.

4. If required, select the Java options to be uninstalled.

Note:
These options should only be selected with care, as they may be required by other software
installed on the User’s PC.

5. Select the Next > button and follow the on-screen instructions.

Note:
1. In Windows 2000, the procedure may be initiated via the Control Panel, Add/Remove
Programs dialog. Select MassLynx V4 in the list box and select the Remove button; a
prompt will appear, asking for the MassLynx Installation CD disk. Insert the disk in the CD-
ROM drive and follow the above procedure.

2. The uninstall process does not remove MassLynx project files. If MassLynx is reinstalled,
these will be detected as a previous MassLynx installation; when prompted, allow the
Installation Wizard to rename their location.

Page 2-10
 Installing MassLynx

Figure 2.7 The Uninstall Option: Uninstall Java components dialog

Repairing MassLynx
On Windows 2000 and Windows XP, MassLynx has the ability to self-repair itself automatically
in the event of MassLynx components being deleted or corrupted. If such a situation occurs the
following message will appear:

Figure 2.8 Auto repair message

The install will attempt to rectify any problems automatically and reinstall files if necessary.

A repair option can also be accessed, as shown in the following procedure, if the MassLynx
program ceases to work correctly. This procedure will fix missing or corrupt files, shortcuts and
registry entries.

1. Insert the MassLynx Installation CD disk into the CD-ROM drive; the installation should start
automatically and invoke the MassLynx Wizard Welcome Screen.

2. Select the Next > button; the Program Maintenance dialog is invoked, see Figure 2.6.

3. Select the Repair option.

4. Select the Next > button and follow the on-screen instructions.

Page 2-11
Installing MassLynx

Note:
In Windows 2000 and Windows XP, the procedure may be initiated via the Control Panel,
Add/Remove Programs dialog. Select MassLynx V4 in the list box and select the Change
button; a prompt will appear, asking for the MassLynx Installation CD disk. Insert the disk in the
CD-ROM drive and follow the above procedure.

Page 2-12
 The MassLynx Window and Related Information

Chapter 3 The MassLynx Window

and Related Information

Page 3-1
The MassLynx Window and Related Information

Contents
Opening MassLynx ....................................................................................................................... 3-5
Closing MassLynx......................................................................................................................... 3-5
The MassLynx Window ................................................................................................................ 3-6
The MassLynx Menu Bar.............................................................................................................. 3-7
General............................................................................................................................... 3-7
The File Menu.................................................................................................................... 3-7
The View Menu ................................................................................................................. 3-9
The Run Menu ................................................................................................................... 3-9
The Security Menu............................................................................................................. 3-9
The Help Menu ................................................................................................................ 3-10
The MassLynx Tool Bar.............................................................................................................. 3-10
General............................................................................................................................. 3-10
The Sample List Menu Bar.......................................................................................................... 3-11
The Sample List........................................................................................................................... 3-11
The MassLynx Bar ...................................................................................................................... 3-11
General............................................................................................................................. 3-11
Displaying the MassLynx Bar.......................................................................................... 3-12
The Shortcut Bar .............................................................................................................. 3-12
The Queue Bar ................................................................................................................. 3-16
The MS Status Bar ........................................................................................................... 3-17
Changing the Colors and Fonts in MassLynx Windows ............................................................. 3-18
The Colors and Fonts dialog ............................................................................................ 3-18
The Font Dialog ............................................................................................................... 3-19
The Color Dialog ............................................................................................................. 3-20
The Define Custom Color Dialog .................................................................................... 3-21
To Change MassLynx Fonts or Colors............................................................................. 3-21
MassLynx System Global Parameters ......................................................................................... 3-22
General............................................................................................................................. 3-22
The Options Dialog .......................................................................................................... 3-22
The Options, Multi-probe Dialog..................................................................................... 3-25
The Masslynx.ini File.................................................................................................................. 3-26
General............................................................................................................................. 3-26
To Restore Masslynx.sav Using the Windows Explorer.................................................. 3-26
Selecting and Viewing Data ........................................................................................................ 3-27
General............................................................................................................................. 3-27
Opening Data Files: The MassLynx Window Data Browser Dialog ............................... 3-27
The History Selector Dialog............................................................................................. 3-28
The Experimental Record Window.................................................................................. 3-31
Using Windows Explorer to Open Multiple Data Files............................................................... 3-32
Projects ........................................................................................................................................ 3-33
General............................................................................................................................. 3-33
To Create a New Project .................................................................................................. 3-33
To Open an Existing Project ............................................................................................ 3-35
Directory Structure ...................................................................................................................... 3-35
Data File Structure ........................................................................................................... 3-37
Displaying Spectra ...................................................................................................................... 3-37
To Display a Spectrum Using the Sample List Menu Bar ............................................... 3-37
To Display a Spectrum from a Chromatogram ................................................................ 3-37
To Remove Spectra and Spectrum Windows................................................................... 3-38
Displaying Chromatograms......................................................................................................... 3-38
To Display a Chromatogram Using the Sample List Menu Bar....................................... 3-38
To Display a Chromatogram from Spectrum ................................................................... 3-38
To Remove Chromatogram Traces and Chromatogram Windows .................................. 3-38
The Header Editor Dialog ........................................................................................................... 3-39
General............................................................................................................................. 3-39
The User Text Dialog....................................................................................................... 3-40
The Format Dialog ........................................................................................................... 3-41

Page 3-2
 The MassLynx Window and Related Information

To Add Information to the Displayed Header in a MassLynx Program Window............ 3-41

To Remove Information from the Displayed Header....................................................... 3-42
Printing Data ............................................................................................................................... 3-42
General............................................................................................................................. 3-42
Printing a Specific MassLynx Window Using the Tool Bar............................................ 3-43
Printing a Specific MassLynx Window Using the Menu Bar .......................................... 3-43
Window Commands.................................................................................................................... 3-44
Getting Help................................................................................................................................ 3-44

Illustrations
Figure 3.1 The MassLynx Login Window ................................................................................... 3-5
Figure 3.2 Typical MassLynx Window........................................................................................ 3-6
Figure 3.3 The File Menu............................................................................................................. 3-7
Figure 3.4 Typical sample List Properties Window ..................................................................... 3-8
Figure 3.5 The View Menu .......................................................................................................... 3-9
Figure 3.6 The Run Menu ............................................................................................................ 3-9
Figure 3.7 The Security Menu...................................................................................................... 3-9
Figure 3.8 The Import Project sub-menu ................................................................................... 3-10
Figure 3.9 The Help Menu ......................................................................................................... 3-10
Figure 3.10 The View, MassLynx Bar Menu............................................................................. 3-12
Figure 3.11 The Instrument Shortcut Bar................................................................................... 3-13
Figure 3.12 The Tools Shortcut Bar........................................................................................... 3-15
Figure 3.13 Typical Application Manager Shortcut Bar ............................................................ 3-16
Figure 3.14 Typical MS Status Bar ............................................................................................ 3-17
Figure 3.15 The Colors and Fonts dialog ................................................................................... 3-18
Figure 3.16 The Font Dialog ...................................................................................................... 3-19
Figure 3.17 The Color dialog ..................................................................................................... 3-20
Figure 3.18 The Define Custom Color dialog ............................................................................ 3-21
Figure 3.19 The Options dialog.................................................................................................. 3-23
Figure 3.20 The Options, Multi-probe dialog ............................................................................ 3-25
Figure 3.21 The MassLynx Window Data Browser Dialog ....................................................... 3-27
Figure 3.22 The History Selector dialog .................................................................................... 3-29
Figure 3.23 The Experimental Record Window......................................................................... 3-31
Figure 3.24 The Experimental Record window File menu ........................................................ 3-32
Figure 3.25 The Experimental Record window Options menu .................................................. 3-32
Figure 3.26 Project warning dialog ............................................................................................ 3-33
Figure 3.27 The Create Project dialog (1) .................................................................................. 3-34
Figure 3.28 The Create Project dialog (2) .................................................................................. 3-34
Figure 3.29 The Select Project dialog ........................................................................................ 3-35
Figure 3.30 The Header Editor dialog........................................................................................ 3-39
Figure 3.31 The User Text dialog .............................................................................................. 3-40
Figure 3.32 The Format dialog................................................................................................... 3-41
Figure 3.33 The Print dialog ...................................................................................................... 3-43

Page 3-3
The MassLynx Window and Related Information

Page 3-4
 The MassLynx Window and Related Information

Opening MassLynx
Either:

1. a) Select the Windows Start button; the Start menu is invoked.

b) Select the Programs, MassLynx, MassLynx V4.0 option.

Or:

2. Double-click on the MassLynx V4.0 icon on the Windows desktop.

In either case, if MassLynx Security is not enabled, MassLynx will start and the MassLynx
Window will appear, see the “The MassLynx Window” section, on page 3-6. If MassLynx
Security is enabled, the MassLynx Login window will be displayed.

Figure 3.1 The MassLynx Login Window

3. Enter the Logon Name:, Password: and Domain.

4. Select the OK button.

After a few seconds, MassLynx will start and the MassLynx Window will appear, see the “The
MassLynx Window” section, on page 3-6.

If problems are experienced, these may be due to the security set up, see the “MassLynx Security
User’s Guide” for further information.

Closing MassLynx
A MassLynx session is terminated in the normal Windows way, either by clicking on the windows
close box, at the top right-hand corner of the MassLynx Window, or by selecting the Menu Bar
File, Exit command.

If Windows is to be shutdown while MassLynx is running, MassLynx will display a message box
asking if MassLynx is to be shut down. If the OK button is selected, MassLynx will terminate

Page 3-5
The MassLynx Window and Related Information

followed by Windows; if the Cancel button is selected, both MassLynx and Windows will
continue running.

If data acquisition is in progress and Windows identification is requested to shutdown, MassLynx

will warn that data will be lost if it is terminated and ask if should still continue. If the Yes button
is selected, the acquisition will stop and Windows will be closed; if the Cancel button is selected,
data acquisition will continue.

The MassLynx Window

The MassLynx Window is invoked when MassLynx is started.

Banner Menu Bar Tool Bar Information Bar Sample List Menu Bar

Status Bar MassLynx Bar Sample List Editor

Figure 3.2 Typical MassLynx Window

The Window contains:

• A Banner, which displays the names of the current Project and Sample List.

• A top-level Menu Bar, see the “The MassLynx Menu Bar” section, on page 3-7.

• A Tool Bar, see the “The MassLynx Tool Bar” section, on page 3-10.

• An Information Bar (below the Tool Bar); this normally shows the status of the sample
currently being acquired or processed.

Page 3-6
 The MassLynx Window and Related Information

• The MassLynx Bar, with associated tabs, at the left-hand-side of the window; see the “The
MassLynx Bar” section, on page 3-11. The contents of the MassLynx Bar can be swapped
between the Shortcut Bar, Queue Bar and Instrument Status Bar, each of which has its own
set of associated tabs and options.

• The Sample List Editor, see Chapter 4, “Sample Lists”.

• A Sample List Menu Bar (above the Sample List Editor), containing commands associated
with the Sample List; see the “The Sample List Menu Bar” section, on page 3-11.

• A Status Bar, at the bottom of the window.

Multiple windows, such as Chromatogram or Spectrum displays, can be invoked in the MassLynx
Window as required; these windows may be moved around the MassLynx Window and, in certain
cases, resized.

The MassLynx Menu Bar

General

The MassLynx Menu Bar appears at the top of the MassLynx Window. The Menu Bar gives
access to the facilities used to customize the MassLynx Window, control projects, data files, etc.
and control data acquisition.

The File Menu

Figure 3.3 The File Menu

Page 3-7
The MassLynx Window and Related Information

Open Project Opens an existing project, see the “To Open an Existing Project” section,
on page 3-35.

Project Wizard Creates a new project, using the Project Wizard, see the “To Create a New
Project” section, on page 3-33.

Open Data File Opens an existing data file, see the “Opening Data Files: The MassLynx
Window Data Browser Dialog” section, on page 3-27.

New Creates a new Sample List, see Chapter 4, “Sample Lists”.

Open Opens an existing Sample List, see Chapter 4, “Sample Lists”.

Save Saves the current Sample List to disk, see Chapter 4, “Sample Lists”.

Save As Saves a copy of the current Sample List to disk with a new file name, see
Chapter 4, “Sample Lists”.

Sign Sample List Allows the Sample List to be signed under “Secure Files” conditions.

Note:
This option is only displayed when MassLynx has been installed with the
“Secure Files” security option; see the “MassLynx Security User’s
Guide” for details.

Sample List Invokes a window giving details of the current Sample List; details include
Properties the file name and location, and when the file was last modified.

Figure 3.4 Typical sample List Properties Window

Import Imports a worksheet. The worksheet can be an OpenLynx batch file, a tab
Worksheet delimited text file, a comma separated text file, an Excel spreadsheet, or an
Access 97 file, see Chapter 4, “Sample Lists”.

Import Data Imports spreadsheet/database information into the Sample List Editor.
Formats supported are: Excel 5.0/Excel 97 (*.xls), Access 97 (*.mdb,
*.spl), tab delimited text (*.tdl, *.tdb, *.txt) and comma delimited text
(*.cvs, *.txt) , see Chapter 4, “Sample Lists”.

Print Prints the Sample List, see Chapter 4, “Sample Lists”.

Page 3-8
 The MassLynx Window and Related Information

Print Setup Selects the printer to be used via the standard Windows Printer Setup
dialog.

Exit Closes the MassLynx application.

The View Menu

Figure 3.5 The View Menu

Toolbar Toggles the Tool Bar on and off.

Status Bar Toggles the Status Bar on and off.

MassLynx Bar Allows the MassLynx Bar display to be selected; see the “Displaying the
MassLynx Bar” section, on page 3-12 for details.

The Run Menu

Figure 3.6 The Run Menu

Start Starts data acquisition, see Chapter 5, “The MassLynx Queue”.

Stop Stops data acquisition for the currently running job.

Pause Pauses data acquisition for the currently running job.

The Security Menu

Figure 3.7 The Security Menu

Note:
The Security Menu is only displayed when MassLynx security is enabled.

Lock MassLynx Locks MassLynx; to use MassLynx the User must log in again.

Log Off Logs the current User off MassLynx; a new User can then log in.

Page 3-9
The MassLynx Window and Related Information

Secure File Invokes the Import Project sub-menu.

System
Import Project Allows a full project directory of legacy files to be imported under “Secure
Files” conditions.

Note:
This option is only displayed when MassLynx has been installed with the
“Secure Files” security option; see the “MassLynx Security User’s
Guide” for details.

Figure 3.8 The Import Project sub-menu

The Help Menu

Figure 3.9 The Help Menu

Contents and Opens the Help file; this provides on-line information about using the
Index MassLynx application.

About MassLynx Displays the About MassLynx box, which provides information about
MassLynx, including the version number.

The MassLynx Tool Bar

General

The MassLynx Tool Bar is at the top of the MassLynx Window, below the Menu Bar; it is used to
perform common operations with a single click of the appropriate mouse button. To see what
operation a Tool Bar button performs, move the mouse pointer over the button and a description
will appear.

Tool Bar Menu equivalent Purpose

button
File, Open Project Opens an existing project, see the “To Open an
Existing Project” section, on page 3-35. Selecting

the arrow, , displays a list of the most recent

projects.
File, New Creates a new Sample List, see Chapter 4, “Sample
Lists”.

Page 3-10
 The MassLynx Window and Related Information

Tool Bar Menu equivalent Purpose

button
File, Open Opens an existing Sample List, see Chapter 4,
“Sample Lists”.
File, Save or Saves a Sample List, see Chapter 4, “Sample Lists”.
File, Save As
File, Print Prints the Sample List, see Chapter 4, “Sample
Lists”.
Run, Start Starts data acquisition, see Chapter 5, “The
MassLynx Queue”.
Run, Stop Stops data acquisition.

Run, Pause Pauses data acquisition.

View, MassLynx Bar, Invokes the Shortcut Bar; this Bar can be changed
Shortcut to display functions associated with the Instrument,
Tools and MassLynx-associated Application
Managers, such as QuanLynx, BioLynx, etc. See
the “The Shortcut Bar” section, on page 3-12 for
details.
View, MassLynx Bar, Invokes the Queue Bar; this is used to control the
Queue Queue; see the “The Queue Bar” section, on
page 3-16.
View, MassLynx Bar, Invokes the MS Status Bar; see the “The MS Status
Instrument Status Bar” section, on page 3-17.
Security, Lock Locks MassLynx; to use MassLynx the User must
MassLynx log in again.
Note:
This tool is only displayed when MassLynx security
is enabled.

The Sample List Menu Bar

The Sample List Menu Bar contains commands, and two menus, associated with the Sample List;
refer to Chapter 4, “Sample Lists” for further information.

The Sample List

The MassLynx Sample List is a list of the samples available for analysis by the mass spectrometer;
refer to Chapter 4, “Sample Lists” for further information.

The MassLynx Bar

General

The MassLynx Bar is displayed at the left-hand-side of the MassLynx Window. It is a multi-
function display that can be swapped between the Shortcut Bar, Queue Bar and Instrument
Status Bar. Each of these has its own set of associated tabs and options. In turn, the Shortcut Bar

Page 3-11
The MassLynx Window and Related Information

can be swapped between the Instrument Bar, Tools Bar and any installed Application Manager
Bar, as required.

Displaying the MassLynx Bar

The MassLynx Bar displayed is selected via the Menu Bar View, MassLynx Bar menu, or the
appropriate Tool Bar button.

Figure 3.10 The View, MassLynx Bar Menu

Shortcut Selects the Shortcut MassLynx Bar; this Bar provides shortcuts to
functions associated with the Instrument, Tools and MassLynx Application
Managers, such as QuanLynx, BioLynx, etc., by selecting the appropriate
tab from the tabs on the left-hand-side of the Bar. See the “The Shortcut
Bar” section, on page 3-12 for details.

Queue Selects the Queue MassLynx Bar; this is used to control the Queue. See
the “The Queue Bar” section, on page 3-16.

Instrument Selects the MS Status MassLynx Bar; see the “The MS Status Bar”
Status section, on page 3-17.

Note:
If none of the MassLynx Bar options is selected, the MassLynx Bar will be hidden.

The Shortcut Bar

General

The Shortcut Bar is invoked by selecting the Menu Bar View, MassLynx Bar, Shortcut

command, or the Tool Bar button. This Bar provides shortcuts to functions
associated with the Instrument, Tools and MassLynx Application Managers, such as QuanLynx,
BioLynx, etc., by selecting the appropriate tab from the tabs on the left-hand-side of the Bar. To

view additional tabs, click on the symbol.

A question mark symbol ( ) appears on certain Shortcut Bar title graphics; clicking on it will
invoke the Help application for that particular group of applications.

The Instrument Shortcut Bar

The Instrument Shortcut Bar is invoked by selecting the Instrument tab on the left-hand-side of
the Shortcut Bar.

The icons in the Bar are used to select functions associated with the Instrument; to view additional

icons, click on the and symbols, as required.

Page 3-12
 The MassLynx Window and Related Information

Figure 3.11 The Instrument Shortcut Bar

Available icons are:

Control Panel Invokes the Acquisition Control Panel dialog; this accesses and manages
all MassLynx Acquisition functions. Refer to the appropriate Instrument
User’s Guide for details.

Note:
This icon is only present when an instrument with a TDAT interface is
installed.

Inlet Method Invokes the Inlet Method dialog; this is used to view the status of the
current system, change instrument configuration, define the autosampler
and detector methods, control pumps, control indicators and run methods.
Refer to the “MassLynx Inlet Control Guide ” for details.

Page 3-13
The MassLynx Window and Related Information

MS Method Invokes the MS Method Editor; this is used to set up the scanning
function(s) used to scan the instrument during an acquisition. Refer to the
appropriate Instrument User’s Guide for details.

Note:
This icon is only present if a mass spectrometer has been specified during
MassLynx installation.

MS Tune Invokes the Tune Page; this is used to modify the instrument tuning
parameters. Refer to the appropriate Instrument User’s Guide for details.

Note:
This icon is only present if a mass spectrometer has been specified during
MassLynx installation.

Edit Shutdown Invokes the Shutdown Editor; this is used to edit the automatic startup and
or Startup shutdown files, or to create new files. Refer to the appropriate Instrument
User’s Guide for details.

Shutdown Runs the automatic Shutdown file. Refer to the appropriate Instrument
User’s Guide for details.

Startup Runs the machine-specific automatic Start-up file; once completed the
instrument will be in a condition to acquire data. Refer to the appropriate
Instrument User’s Guide for details.

Options Invokes the Options, Multi-probe dialog; this is used edit the instrument
probe parameters, see the “The Options, Multi-probe” section, on
page 3-25, for details.

The Tools Shortcut Bar

The Tools Shortcut Bar is invoked by selecting the Tools tab on the left-hand-side of the Shortcut
Bar.

The icons in the Bar are used to select MassLynx tools; to view additional icons, click on the

and symbols, as required.

Available icons are:

Options Invokes the Options dialog; see the “The Options Dialog” section on
page 3-22.

Colors and Fonts Invokes the Colors and Fonts dialog; refer to the “Changing the Colors and
Fonts in MassLynx Windows” section, on page 3-18.

Strip Invokes the Strip Datafile dialog; the Strip utility removes unwanted
background and noise from a data file. Refer to Chapter 8, “Strip and
Combine Functions” for details.

Page 3-14
 The MassLynx Window and Related Information

Figure 3.12 The Tools Shortcut Bar

Accurate Mass Invokes the Accurate Mass Measure (AMM) dialog; this utility provides
Measure a variety of post acquisition Mass Measure data processing facilities that
can be applied to whole files. Refer to Chapter 15, “Accurate Mass
Measure” for details.

Combine Invokes the Combine Datafile Functions dialog; this utility provides a
Functions way of combining all the functions in a data file to produce a new data file
containing a single function, which is the sum of the multiple functions.
Refer to Chapter 8, “Strip and Combine Functions” for details.

Combine All Combines a group of files that have all been acquired using the same
Files acquisition method to produce a single output file. This results in an
increase in the signal to noise ratio. Refer to Chapter 8, “Strip and
Combine Functions” for details.

Search Library Invokes the Library – [Hits] Window; refer to Chapter 11, “Library” for
details.

Page 3-15
The MassLynx Window and Related Information

Molecular Invokes the Molecular Mass Calculator dialog; refer to Chapter 12,
Weight “Molecular Mass Calculator” for details.
Calculator
Application Managers Shortcut Bars

Each installed MassLynx Application Manager has its own Shortcut Bar, which is invoked by
selecting the appropriate tab on the left-hand-side of the Shortcut MassLynx Bar. A typical
Application Manager Shortcut Bar is shown below. Refer to the appropriate User’s Guide for the
Application for full details.

Figure 3.13 Typical Application Manager Shortcut Bar

The Queue Bar

The Queue Bar is invoked by selecting the Menu Bar View, MassLynx Bar, Queue command, or

the Tool Bar button. This Bar is used to manage the MassLynx Queue; refer to
Chapter 5, “The MassLynx Queue” for details.

Page 3-16
 The MassLynx Window and Related Information

The MS Status Bar

The MS Status Bar is invoked by selecting the Menu Bar View, MassLynx Bar, Instrument

Status command, or the Tool Bar button.

Figure 3.14 Typical MS Status Bar

The MS Status Bar shows the current instrument and inlet status information. Three tabs are on
the left-hand-side of the MS Status Bar:

Inlet Method Invokes the Inlet Method dialog; refer to the “MassLynx NT Guide to Inlet
Control” for details.

MS Method Invokes the MS Method Editor; refer to the appropriate instrument User’s
Guide for details.

Note:
This icon is only present if a mass spectrometer has been specified during
MassLynx installation.

Page 3-17
The MassLynx Window and Related Information

Tune Invokes the Tune dialog; refer to the appropriate instrument User’s Guide
for details.

Note:
This icon is only present if a mass spectrometer has been specified during
MassLynx installation.

Changing the Colors and Fonts in MassLynx

Windows
The Colors and Fonts dialog

The fonts and colors used in MassLynx windows can be altered to suit the User’s preferences
using the Colors and Fonts dialog. This dialog is invoked by selecting the Tools Shortcut Bar,
Colors and Fonts icon, see the “The Tools Shortcut Bar” section, on page 3-14.

Figure 3.15 The Colors and Fonts dialog

Type: Frame The list box details items for which the colors/fonts can be changed; the
item for which color/font is to be changed can be selected by clicking on
it. Double-clicking on an item in this Frame will invoke the Color or Font
dialog, as appropriate.

Color Invokes the Color dialog; this allows the item’s color to be changed, see
the “The Color Dialog” section, on page 3-20 for details. This button is
enabled when a non-text item is-selected in the list box.

Font Invokes the Font dialog; this allows the text font and color to be changed,
see the “The Font Dialog” section, on page 3-19 for details. The Font
button is enabled when a text item is-selected in the list box.

Page 3-18
 The MassLynx Window and Related Information

Current Settings Displays the fonts and colors currently in use. Double-clicking on an item
Frame in this Frame will invoke the Color or Font dialog, as appropriate.

Note:
1. Data colors 1 to 5 are used for Chromatogram and Spectrum
displays.

2. Data color 5 is also used to set the color of tune peaks in the Tune
Page.

3. Data colors 6 to 10 are used for the fill colors on peak detected
chromatograms, components in ElectroSpray spectra and for the Map
program.

The Font Dialog

The Font Dialog is invoked as shown in the “Changing the Colors and Fonts in MassLynx
Windows” section, on page 3-18.

Figure 3.16 The Font Dialog

Font: Select the required font from the list, or type its name in the text box.

Font style: Select the required font style from the list, or type its name in the text box.

Size: Select the required font size from the list, or type a value in the text box.

Effects Frame

Strikeout Strikes out the text.

Page 3-19
The MassLynx Window and Related Information

Underline Underlines the text.

Color: Select the required text color from the drop-down list.

Sample Displays an example of text formatted in accordance with the selections in

this dialog.

Script: Select the required script from the drop-down list.

OK Closes the Font dialog and returns to the Colors and Fonts dialog.

The Color Dialog

The Color dialog is invoked as shown in the “Changing the Colors and Fonts” section, on
page 3-18.

Figure 3.17 The Color dialog

Basic colors: Forty-eight basic colors are displayed; click on the required color to select
it.

Custom colors: Displays colors created in the Define Custom Color dialog, see the “The
Define Custom Color Dialog” section, on page 3-21.

Define Custom Invokes the Define Custom Color dialog, see the “The Define Custom
Colors>> Color Dialog” section, on page 3-21.

OK Closes the Color dialog and returns to the Colors and Fonts dialog.

Page 3-20
 The MassLynx Window and Related Information

The Define Custom Color Dialog

The Define Custom Color dialog is invoked by the Color dialog, Define Custom Colors>>
button.

Figure 3.18 The Define Custom Color dialog

To define custom colors:

1. Either:

Drag the cross-hairs and the arrow until the required color appears in the Color|Solid
box.

Or:

Type values into the Hue:, Sat:, Lum:, Red:, Green: and Blue boxes until the required color
appears in the Color|Solid box.

2. Select the Add to Custom Colors button. The new color will appear in the next available
Custom colors box on the left of the dialog.

3. Select the OK button. The Define Custom Color dialog is closed and the display returns to
the Color Dialog.

To Change MassLynx Fonts or Colors

1. Select the Tools Shortcut Bar, Colors and Fonts icon; the Colors and Fonts dialog is
invoked.

2. Either:

a. Select the required item in the Type: Frame. The Color or Font button will be enabled,
depending on the type of item selected.

b. Select the Font or Color button, as appropriate.

Page 3-21
The MassLynx Window and Related Information

Or:

Double-click on the required item in the Type: Frame.

Or:

Double-click on the required item in the Current Settings Frame.

In any of the above cases, the Font or Color dialog is invoked as appropriate; see the “The
Font Dialog” section, on page 3-19, and the “The Color Dialog” section, on page 3-20, for
details.

3. Make the required changes in the Font or Color dialog before selecting the OK button. The
Font or Color dialog is closed and the revised details are displayed in the Colors and Fonts
dialog Current Settings Frame.

4. Repeat steps 2 and 3 for each item to be changed.

5. Select the OK button. The changes are saved and the Colors and Fonts dialog is closed.

MassLynx System Global Parameters

General

There are various MassLynx parameters that need to be applied to several windows in the system,
such as whether to work in retention times or scans. Rather than setting these controls in every
window, these values are set once at the top level, and they are then used, where relevant, within
the rest of the system. These are the System Global Parameters and they can be modified as
required. For system-related parameters, refer to the “The Options Dialog” section, below; for
instrument-related parameters, refer to the “The Options, Multi-probe Dialog” section, on
page 3-25.

The Options Dialog

The Options dialog is invoked by selecting the Tools Shortcut Bar, Options icon, see “The Tools
Shortcut Bar” section, on page 3-14.

Display Type Specifies whether the units for spectra and chromatograms are to be scan
Frame numbers or retention times.

Scan Number When selected, and the Spectrum or Library Tool Bar button is
pressed, the required value will be in the Scan Number format.

Retention Time When selected, and the Spectrum or Library Tool Bar button is
pressed, the required value will be in the Retention Time format.

Page 3-22
 The MassLynx Window and Related Information

Figure 3.19 The Options dialog

Mass
Chromatogram
Window Frame
Parts per Displays the Chromatogram Window in parts per million. For accurate
million mass chromatograms, Parts per million should be selected with a value of
between 5 and 10 entered in the adjacent text box.

Abs window Should be selected for data other than mass chromatograms; for
(Da) quadrupole data the default value of 1 should be entered in the adjacent
text box. For magnetic sector or Tof (time-of-flight) data, the
Abs window (Da) value may need to be decreased.

Raw data Frame Determines whether to display TIC (Total Ion Current) or Mass
Chromatograms for data acquired using the MUX (multi-injector) system.

Display
chromatogram
as:
TIC Displays TIC (Total Ion Current) chromatograms.

Page 3-23
The MassLynx Window and Related Information

Mass When selected and there is a mass in any of the Mass A to Mass T
Chromatogram columns in the Sample List (see Chapter 4, “Sample Lists”), the Mass
Chromatogram is displayed, otherwise the TIC is displayed.

Adducts ion
mass:
Positive ion and If a value is entered in either of these boxes, the MassLynx software will
Negative ion automatically apply the correct adduct depending on the ion mode of the
data file being displayed.

Axes Labelling: Determines axis labeling for spectral displays and can be chosen from
Da/e, u/e or m/z where:

Da represents Daltons.

u or m represents atomic mass units.

e or z represents the elementary charge.

Note:
This labeling will not apply to ElectroSpray spectra that have been
transformed onto a true molecular mass scale.

Use Acquired Select this option to always use the last acquired raw data file when the
File as Default Spectrum or Chromatogram Windows are initialized.

MassLynx Status Specifies whether to save the MassLynx system state, how often and to
Frame which file.

Update Status Select this option to write the status of the instrument to a file. The default
file is c:\masslynx\status.ini; to change this, select the File Name button.
These instrument status files can be viewed in a text editor, such as
Notepad, across a network, allowing Users to decide which instrument
should be used to acquire samples. Each file will contain the MS status,
the LC status and details of samples in the Queue.

Refresh rate By default, the details in the instrument status file are updated every
60 seconds; to change this, enter a new time in this box.

File Name Invokes a browser that allows the current instrument status file to be
changed to another file. The current instrument status file is displayed in
the adjacent box.

Database Specifies whether to save each sample, and to which file.

Logging Frame
Log Samples Select this option to write details of all samples acquired to a database file.
The default database file is c:\masslynx\sample.mdb. The database can be
used to analyze machine usage.

Database Invokes a browser that allows the default database file to be changed. The
current database file is displayed in the adjacent box.

Page 3-24
 The MassLynx Window and Related Information

Note:
If the settings in the MassLynx Status or the Database Logging Frames are changed, MassLynx
must be restarted for the changes to take effect.

The Options, Multi-probe Dialog

The Options, Multi-probe dialog is invoked by selecting the Instrument Shortcut Bar, Options
icon, see “The Instrument Shortcut Bar” section, on page 3-12.

Figure 3.20 The Options, Multi-probe dialog

Multi-probe
Source Frame
Multi-probe Select this option when the instrument being used has multi-probe
Capability capability.

Note:
Selecting the Multi-probe Capability option disables the Dual Source, Lock Spray option.

Probes Enter the number of probes in use in this text box.

Accurate Mass Select this option to use the accurate mass facility.

Pos/Neg Select this option to indicate the use of positive/negative switching.

Switching

Dual Source, Select this option to use a dual source lock spray.
Lock Spray

Page 3-25
The MassLynx Window and Related Information

Note:
Selecting the Dual Source, Lock Spray option disables the Multi-probe Source Frame.

Inlets Frame

Allow Random Allows random bottle locations to be used in a Sample List.

Bottle
Locations in
Sample List
Multi-Inlet Allows multiple inlets to be used; the number of inlets is entered in the
Capability Number of Parallel Inlets text box.

Sample Prep Allows multiple inlets to be used on a non-MUX system, using a prep file;
see the “MassLynx Inlet Control Guide” for further information.

The Masslynx.ini File

General

The masslynx.ini file contains current settings for all MassLynx windows and dialog boxes. If the
MassLynx Security Manager Menu Bar Policies, Use Individual INI files command is selected
(see the “MassLynx Security User’s Guide”), when a new User logs to on to MassLynx, a new
username.ini file is created. Each time this User uses MassLynx any changes to the current
settings are saved to this file. It is possible that, under some conditions, the settings in
username.ini may become corrupted and cause problems with the operation of MassLynx. A
default .ini file is stored in the c:\masslynx directory, saved under the name masslynx.sav. This
backup file can be copied to username.ini to restore a set of uncorrupted default parameters.

To Restore Masslynx.sav Using the Windows Explorer

1. Save any parameter files that are needed in MassLynx, e.g. tuning parameter files.

2. Close down MassLynx.

3. Open the Windows Explorer.

4. Navigate to the MassLynx directory.

5. Select the masslynx.sav file.

6. Select the Edit, Copy command.

7. Select the Edit, Paste command; this will produce a file called “copy of masslynx.sav”.

8. Delete your username.ini file, e.g. administrator.ini.

9. Rename the “copy of masslynx.sav” file the your username.ini.

10. Restart MassLynx.

11. This will set the MassLynx system, for this User, back to the default state.

Page 3-26
 The MassLynx Window and Related Information

Selecting and Viewing Data

General

This section deals with the basic procedures for selecting and displaying data. More detailed
information is provided later in the manual.

Opening Data Files: The MassLynx Window Data Browser Dialog

Data files are opened using the Data Browser dialog; this is invoked by the Menu Bar File, Open
Data File command.

Note:
The Spectrum, Chromatogram and Library functions each have their own Data Browser
dialogs; refer to the appropriate sections of this User’s Guide for details.

Figure 3.21 The MassLynx Window Data Browser Dialog

File Name: The required file name may be typed in the text box, or one may be
selected from the list below by clicking on it. The file name may include a
path, if required.

Directories: Lists the directories available on the current drive.

Page 3-27
The MassLynx Window and Related Information

Drives: Lists the other available drives.

Network Invokes the Windows Map Network Drive dialog, so that a connection can
be made to a shared network directory.

Information Contains information relevant to the currently highlighted data file.

Frame
Sample Displays the sample description, obtained from the header of the currently
Description selected data file. This is information, such as compound name and
concentration, which was entered during acquisition.

Acquired Displays the date and time at which this file was acquired.

Function Displays the currently selected scan function. The function description
gives the function type, mass range and ionization mode. A new function
can be selected from the drop down list box.

History Invokes the History Selector dialog, which provides access to processed
data; see the “The History Selector Dialog” section, below, for details.

Experiment Invokes the Experimental Record dialog; this displays information about
the raw data file, see the “The Experimental Record Window” section, on
page 3-29, for details.

Delete Deletes the selected raw data file.

Chromatogram Automatically loads the Chromatogram window, displaying the

Chromatogram of the new data file, when the OK button is selected.

Spectrum Automatically loads the Spectrum window, displaying the spectrum of the
new data file, when the OK button is selected.

The History Selector Dialog

General

The History Selector dialog is invoked by the MassLynx Window Data Browser dialog History
button; it provides access to processed data. If no processed data is selected, raw data is the
default. When raw data is processed (for example, using the Refine or Combine functions), the
processed data can be saved using the Spectrum File, Save Spectrum command.

The History Selector dialog also allows the deletion of processes that are no longer required.

Process History: Shows the full history of all saved processes with the original raw data at
the top of the tree.

Processed data, which has been derived from previously processed data, is
indented to show its relationship to this data. Each process is labeled with
a unique identification number and the time and date when it was created;
this aids differentiation of similar processes. Refer to the “Processed Data
Labels” section, on page 3-29, for details of the process identification
labels.

Page 3-28
 The MassLynx Window and Related Information

Figure 3.22 The History Selector dialog

Information
Frame
Sample: Displays sample description text obtained from the header of the currently
selected data file.

Function: Displays a description of the currently selected function.

History: Displays full history of the currently selected process. This starts with raw
data at the top of the list and describes each processing step made to reach
the current process.

Note:
Details of the processing step are only displayed in the History: Frame only after they have been
selected in the Process History: window.

OK Exits the History Selector dialog using the current selection.

Cancel Exits the History Selector dialog defaulting to the original selection.

Delete Deletes the currently selected process from the Process History: tree.

Delete All Deletes all processes belonging to the current data file function.

Processed Data Labels

Each of the processed data labels is followed by a series of letters and numbers that describe the
parameters used during the process:

Refine Rf (n1, n2)

Rf = Refined spectrum.

n1 = Refine window in scans.

Page 3-29
The MassLynx Window and Related Information

n2 = Refine noise level.

Combine Cm (n1:n2 - (n3:n4 + n5:n6) x n7)

Cm = Combined spectrum.

n1:n2 = Average range start and end values.

n3:n4 = First subtract range start and end values.

n5:n6 = Second subtract range start and end values.

n7 = Subtract range multiplication factor.

Smooth Sm (s1, [n1x], n2)

Sm = Smoothed data.

s1 = Smooth type. Mn - Mean, Md - Median, Sg - Savitzy Golay.

n1x = Number of smooths (not for median).

n2 = Smooth window. MassLynx requires the User to enter an estimate of the width of the raw
data peak at half height in Daltons, and uses this to calculate the width of the smoothing
window. See Chapter 7, “Spectrum” for the definition of the rule used for this calculation.

Subtract Sb (n1, n2)

Sb = Spectrum which has been baseline subtracted.

n1 = order of polynomial which has been fitted to baseline.

n2 = Percentage of data points which lie below baseline.

Center Cn (s1, n1, [n2], s2)

Cn = Centered data.

s1 = Centering method. Top - Highest point on peak, Med - Median of peak, Cen - Centroid of
peak.

n1 = Peak width at half height.

n2 = Topmost percentage of peak used to calculate centroid.

s2 = method used for calculating peak intensities, height “Ht” or area “Ar”.

Transform Tr (n1:n2, n3, s1)

Tr = Transformed spectrum.

n1 = Raw mass range start.

n2 = Raw mass range end.

n3 = Resolution of transformed spectrum in Da/channel.

s1 = Mid or Low, indicates method used to separate overlapping series.

Page 3-30
 The MassLynx Window and Related Information

MaxEnt ME [Ev n1, It n2] (s1, n3, n4:n5, Ln6, Rn7)

n1 = MaxEnt evidence.

n2 = Number of iterations.

s1 = indicates type of damage model used for MaxEnt reconstruction.

Gs - Constant width Gaussian or Sp - Isotopic model and Mass Spectrometer Blur.

n3 = Width of Gaussian in Da at half height for Gs or Width in Da at half height of Mass

Spectrometer blur.

n4 = Raw mass range start.

n5 = Raw mass range end.

Ln6 = Left minimum intensity ratio.

Rn7 = Right minimum intensity ratio.

MaxEnt Mock Data MK [Ev n1, It n2] (s1, n3, n4:n5, Ln6, Rn7)

Parameters are as for MaxEnt above.

The Experimental Record Window

General

The Experimental Record window is invoked by the MassLynx Window Data Browser dialog
Experiment button; it displays information about the selected raw data file.

Figure 3.23 The Experimental Record Window

Page 3-31
The MassLynx Window and Related Information

Information displayed includes:

• Raw data file header information such as sample description, acquisition date and time, etc.

• Tune parameters.

• Function description.

The Experimental Record Window File Menu

Figure 3.24 The Experimental Record window File menu

Save As Saves the Experimental Record to disk as a text file.

Print Report Prints the displayed Experimental Record; this depends on the options
selected in the Experimental Record Window Options Menu.

Print Setup Invokes the standard Windows Print Setup dialog.

Exit Closes the Experimental Record window.

The Experimental Record Window Options Menu

Figure 3.25 The Experimental Record window Options menu

Header Displays the Header and Instrument Calibration data.

Tune parameters Displays the Instrument Tuning Parameters.

Function Displays the Function data.

Description

Using Windows Explorer to Open Multiple Data

Files
It is also possible to use the Windows Explorer to open several MassLynx data files at once and
display them in Chromatogram or Spectrum.

Open Windows Explorer and the MassLynx Chromatogram or Spectrum program, and arrange the
windows so that both are visible.

Select the MassLynx data files to be viewed in the Explorer window. Several files may be
selected, if required. Then, keeping the left mouse button depressed, drag the files into the

Page 3-32
 The MassLynx Window and Related Information

Chromatogram or Spectrum Window; the Window will be re-displayed showing the first function
in each data file as a separate trace.

Projects
General

MassLynx comes with a number of predefined projects that contain example data. The default
project is where all data is stored until a new project has been selected or created.

All MassLynx data storage is organized into projects. When a MassLynx project is created,
MassLynx creates a new directory, called “project.pro”, and the following sub-directories:

• Acqudb Acquisition settings files

• Curvedb Quantify calibration curves

• Data Raw data files

• Methdb Quantify methods

• Peakdb Peak lists

• Sampledb Sample lists

If a project is created using current or existing project as a template, all files in Acqudb, Methdb
and Sampledb are copied into the new project. If an existing project is not chosen as a template
then all sub-directories will be empty.

Note:
This does not apply to BioLynx data, which will use the same directory structure as the previous
version of MassLynx.

To Create a New Project

1. Select the MassLynx Menu Bar File, Project Wizard command; a project warning dialog is
invoked.

Figure 3.26 Project warning dialog

2. Select Yes to continue.

3. The Create Project dialog is invoked.

Page 3-33
The MassLynx Window and Related Information

Figure 3.27 The Create Project dialog (1)

4. Type a Project name and Description into the appropriate fields.

5. A default Location, for saving the project to, appears in the location box. To save the file to a
different location, type a new one into the box.

6. Select the Next button; the next page of the dialog is invoked.

Figure 3.28 The Create Project dialog (2)

7. Select one of Create new project, Create using current project as template or Create
using existing project as template, as appropriate.

If Create new project is selected, a new project is created, but no method or data files are
created.

If Create using current project as template is selected, a project based on the current
project is created; the method and data files are copied to the new project.

If Create using existing project as template is selected, the Browse button will be enabled;
pressing this button will display the Select existing project dialog, allowing an existing
project to be selected as a template.

8. Select the Finish button; the new project will be created.

Page 3-34
 The MassLynx Window and Related Information

All new data files, sample lists, peak lists, quantify method files and quantify calibration curves
will be saved in this project until the User changes to a new project.

To Open an Existing Project

1. Select the Menu Bar File, Open Project command; a project warning dialog is invoked, see
Figure 3.26.

2. Select Yes to continue; the Select Project dialog is invoked.

Figure 3.29 The Select Project dialog

3. Select the required project and select the OK button; the project is opened.

Directory Structure
When MassLynx is installed, a number of default folders are created, these contain information for
different parts of the program. Files can be opened from and saved to any location the User
specifies, however, MassLynx will look in the default folders for the information first. The
following is a list of folders created in the MassLynx directory.

Folder Name Type of information stored in the folder

Idendb Libraries against which searches are performed. Nist and User-defined
libraries.

Macro Example macro files.

Nucdata Nucleotide sequence data files in the EMBL (European Molecular Biology
Laboratory) standard format.

Nucdb 5’ term, 3’ term and linkage information files, etc.

Page 3-35
The MassLynx Window and Related Information

Folder Name Type of information stored in the folder

Nucembl Nucleotide sequence data files from the Swiss-Prot database.

Note:
Files imported in this format, changed and then saved will be in the
Nucdata folder and format.

Pepdata Protein and peptide sequence data files in the EMBL standard format.

Pepdb C-term, N-term, Digest information files, etc.

Pepembl Protein and peptide sequence data files from the Swiss-Prot database.

Note:
Files imported in this format, changed and then saved will be in the
Pepdata folder and format.

Periodic Periodic table.

Plates Plate layout files for Gilson, Waters 2700 and Waters 2790 autosamplers.

Q-Tof Q-Tof specific run time files.

Racks Bed layout files for Gilson, Waters 2700 and Waters 2790 autosamplers.

Ref Calibration reference files.

Shutdown Shutdown parameters.

Structdb Library structures.

The following table lists the folders that are created within projects:

Folder Name Type of information stored in the folder

Acqudb Acquisition defaults and saved tune page settings, calibrations etc. Inlet
method files.

Curvedb Quantify calibration curve data.

Data Raw data files.

Methdb Method files.

Peakdb Peak list data.

Sampledb Sample Lists.

Page 3-36
 The MassLynx Window and Related Information

Data File Structure

Data acquired from the mass spectrometer are saved into data files on the computer’s hard disk.
These data files may contain more than one acquisition function and may also contain processed
data derived from the original raw data, for example, refined spectra.

All files are acquired to the data directory of the current project.

For example, if the file name is specified as test2, the data files are stored in the directory
c:\MassLynx\currentproject\data\test2. If the data file contains two acquisition functions and two
sets of processed data then the directory listing will be as follows:

_Header.txt Data file header information.

_Funcs.inf Information on functions acquired.

_history.inf Information on how data has been processed.

_expment.inf Experimental record information.

_Func001.dat Data file for first function (one for each function).

_Func001.idx Data file index for first function.

_Func002.dat Data file for second function.

_Func002.idx Data file index for second function.

_proc001.dat First processed data file (one for each process).

_proc001.idx Index for first processed data file.

_proc002.dat Second processed data file.

_proc002.idx Index for second processed data file.

Displaying Spectra
To Display a Spectrum Using the Sample List Menu Bar

Select the Sample List Menu Bar Spectrum command. The Spectrum displayed will be the
current default Spectrum (this will be either the last spectrum viewed, or, if acquisition is in
progress, the last Spectrum acquired). If the Spectrum window is already on display, it becomes
the current window.

Selecting one or more rows (samples) in the Sample List Editor, and then selecting the Sample
List Menu Bar Spectrum command, will invoke Spectrum and attempt to load the data files
associated with the specified samples. This will only work if the data is present on the disk.

To Display a Spectrum from a Chromatogram

Double-click on the Chromatogram at the retention time of interest. The Spectrum closest to that
retention time will be displayed. If the Spectrum window is already on display, the selected
Spectrum will either:

Page 3-37
The MassLynx Window and Related Information

• Be added to the one currently on display,

• Replace the one currently on display (if the Spectrum Tool Bar button is activated),

or,

• Be displayed in a new document window (if the Spectrum Tool Bar button is activated).

To Remove Spectra and Spectrum Windows

To remove a particular Spectrum from a Window, click on the Spectrum and press the Delete key.
There will be a prompt to confirm the deletion; select OK to confirm.

To close a particular Spectrum window, select the Windows close button, , at the top right-hand
corner of the window.

Displaying Chromatograms
To Display a Chromatogram Using the Sample List Menu Bar

Select the Sample List Menu Bar Chromatogram command. The Chromatogram displayed will
be the Total Ion Current (TIC) Chromatogram of the current data file (unless the Options, System
dialog Mass Chromatogram option has been selected and the files selected contain MUX data;
see the “The Options Dialog” section, on page 3-22 for details). If the Chromatogram Window is
already on display, it becomes the current Window.

Selecting one or more rows (samples) in the Sample List Editor, and then selecting the Sample
List Menu Bar Chromatogram command, will invoke Chromatogram and attempt to load the data
files associated with the specified samples. This will only work if the data is present on the disk.

To Display a Chromatogram from Spectrum

Double click on the Spectrum at the mass of interest.

If the Chromatogram Window is already on display, the selected Chromatogram will either:

• Be added to the one currently on display.

• Replace the one currently on display (if the Chromatogram Tool Bar button is activated).

• Be displayed in a new document window of its own (if the Chromatogram Tool Bar
button is activated).

To Remove Chromatogram Traces and Chromatogram Windows

To remove a particular Chromatogram trace, click on the trace and press the Delete key. There
will be a prompt to confirm the deletion; press OK to confirm.

To close a particular Chromatogram Window, select the Windows close button, , at the top
right hand corner of the window.

Page 3-38
 The MassLynx Window and Related Information

The Header Editor Dialog

General

The Header Editor dialog is used to specify the information displayed in the header for each of
the MassLynx program windows. The Header Editor dialog can be invoked from most of the
MassLynx program windows by selecting the Display dialog, Header button (this dialog is
invoked by selecting the Menu Bar Display, View command).

Figure 3.30 The Header Editor dialog

Header areas This is as representation of the MassLynx Window Header, which can be
Frame thought of as a table that has six rows and three columns. Various
information can be displayed in the header, including User-generated text.
Information can be displayed in three positions on each row, left, center, or
right.

Header Editor areas currently displaying information are shaded in gray.

A maximum of eight areas can be used simultaneously.

Cell : Frame A description of the currently-selected header area is displayed at the top
of this Frame.

Group Contains the group name of the list currently displayed in the Element list
box. To change groups, select a different name from the drop-down list.

Element Contains the list of items for the group currently selected in the Group
box. Click on an item to select it.

Format list Displays the items that have been selected from the Element box.

Page 3-39
The MassLynx Window and Related Information

Add -> Adds the currently-selected Element item to the Format list, above the
currently-selected entry. If the currently-selected Element list entry is
[Text], the User Text dialog is invoked; this allows text to be entered in
the Format list box, see the “The User Text Dialog” section, on page 3-40.
This button is grayed if there is no entry selected in the Element list.

Note:
Double-clicking on an Element item will also add it to the Format list

<- Remove Deletes the currently-selected Format list entry. This button is grayed if
there is no entry selected in the Format list.

<- Clear Deletes the entire contents of the Format list for the current cell only.
This button is grayed if there are no entries in the Format list.

Format button Invokes the Format dialog, this allows the currently-selected Element
item to be specified as textual or numeric; see the “The Format Dialog”
section, on page 3-41.

Note:
Double-clicking on an Element item will also invoke the Format dialog.

Clear All Deletes the entire contents of the Format list for all cells. This button is
grayed if there are no entries in the Format list.

The User Text Dialog

The User Text dialog is invoked by the Header Editor, Add -> button, when the currently-
selected Header Editor, Element list entry is [Text]; this allows text to be entered in the Format
list box.

Figure 3.31 The User Text dialog

Text: Text may be typed into this box (41 characters maximum).

OK Closes the dialog and adds the text to the Header Editor, Format list.

Page 3-40
 The MassLynx Window and Related Information

The Format Dialog

The Format dialog is invoked by the Header Editor, Format button, or by double-clicking on an
Element item; this allows the currently-selected information to be specified as textual or numeric.

Figure 3.32 The Format dialog

Type Frame

Textual Marks the item as text.

Numeric Marks the item as a number.

Decimal Places Select the number of decimal places (six maximum) from the drop-down
list. Decimal Places is only available when the Numeric option is
selected.

Field Some elements contain more than one field, for example, directory, file
name, etc. If this is the case, this option is available; select the field to be
displayed from the drop-down list.

To Add Information to the Displayed Header in a MassLynx

Program Window

1. Select the Menu Bar Display, View command; the Display dialog is invoked.

2. Select the Header button; the Header Editor dialog is invoked, see the “General” section, on
page 3-39.

3. Select the required cell in which information is to be displayed in the Header areas Frame.

4. Select the Group that contains the information to be displayed from the drop-down list.

5. Select the required information in the Element list box.

6. To insert information in the Format box, highlight the field before which the information is to
be inserted and select the Add button.

7. To add information at the end of the currently displayed information in the Format box,
highlight End and select the Add button.

Page 3-41
The MassLynx Window and Related Information

8. To add text to the header, select [Text] in the Element list box and select the Add button.
The User Text dialog is invoked, see the “The User Text Dialog” on page 3-40 for details.
Enter the required text and select the OK button. The User text will be shown in the Format
list box and will be displayed in the header when the Header Editor dialog is closed.

9. To format the information in the header, select the relevant field in the Format box and select
the Format button; the Header Editor Format dialog is invoked see the “The Format Dialog”
section, on page 3-41 for details.

10. Repeat steps 1 to 9 as required.

11. Select the OK button to exit and save the changes.

Note:
If the information in one of the Header Editor areas overlaps another area, the overlapped area
will not be displayed.

To Remove Information from the Displayed Header

1. Select the Menu Bar Display, View command; the Display dialog is invoked.

2. Select the Header button; the Header Editor dialog is invoked, see the “General” section, on
page 3-39.

3. Select the required cell from which information is to be removed in the Header areas Frame.

4. To remove a single item, highlight the item to be removed in the Format list box and select
the Remove button.

5. To remove all the information from one Header Editor cell, select the area and select the
Clear button.

6. To remove the information from all the Header Editor cells, select the Clear All button.

7. Repeat steps 1 to 6 as required.

8. Select the OK button to exit and save the changes.

Printing Data
General

MassLynx prints data using the Windows Print Manager.

All of the operations involved in setting up the printer are controlled by Windows and are fully
covered in the “Microsoft Windows NT System Guide”. The only MassLynx specific procedures
are those involved in selecting what to print.

The printer can be set up either using the MassLynx Menu Bar File, Printer Setup command, or
by using the Windows Print Manager.

Page 3-42
 The MassLynx Window and Related Information

Printing a Specific MassLynx Window Using the Tool Bar

Many of the MassLynx Windows have Print buttons on the Tool Bar.

Prints the current Window in portrait format.

Prints the current Window in landscape format.

Printing a Specific MassLynx Window Using the Menu Bar

To print a specific MassLynx window using the Menu Bar commands, select the Window and
select the Menu Bar File, Print command; the Print dialog is invoked.

Figure 3.33 The Print dialog

Print Range
Frame
All Windows Prints all the displayed Windows.

Current Prints the currently selected Window.

Window
Trace Width Select the thickness of the line used to print chromatogram traces or
spectral peaks from the list box; higher values give thicker lines.

Border Prints a border.

Print All Colors Prints the Window in black and white.

Black

Page 3-43
The MassLynx Window and Related Information

Window Commands
Most of the MassLynx program Windows have a Menu Bar Window command. The
subcommands of Window help organize the document Windows so that they fit conveniently into
the main Window. The Window commands are also available on many of the MassLynx Tool
Bars.

Tool Bar Menu Command Function

Button
Tile Arranges open Windows side by side on the screen, dividing
the available space so that all are visible.

To arrange the Windows in a particular order, click on the

title bar of each Window in turn before selecting the Tile
command. The Windows will be tiled in the order in which
they were selected, with the most recently selected Window
first.

Cascade Arranges Windows so that the title bar of each Window is

visible.

Stack Arranges Windows vertically above each other.

Arrange icons Arranges all iconised Windows into rows.

Window list Displays a list of available Windows. The currently active

Window has a tick next to its name. Clicking on another
Window will make that the currently active window. In the
case of Spectrum and Chromatogram this becomes a list
of the traces displayed in each Window.

Window, New Choosing this option causes each subsequent trace to replace
Trace, Replace the currently selected trace.
Trace
Window, New Choosing this option causes each subsequent trace to be
Trace, New displayed in a new Window.
Window
Window, New Choosing this option causes each subsequent trace to be
Trace, Add added to those displayed in the current Window.
Trace

Getting Help
The MassLynx Help system contains detailed information on how to use MassLynx. Most of the
information in this manual is available on-line while using MassLynx by accessing the Help
system.

MassLynx Help can be accessed either from the MassLynx top level Menu Bar or from any of the
MassLynx program windows. It can also be accessed by selecting the MassLynx group
MassLynx User Guide option.

If MassLynx Help is entered from the MassLynx top-level menu, a general index of topics
covering the whole of MassLynx will be invoked. If MassLynx Help is invoked from one of
program windows, help will be given on that particular topic.

Page 3-44
 Sample Lists

Chapter 4 Sample Lists

Page 4-1
Sample Lists

Contents
Introduction ................................................................................................................................... 4-5
The Sample List Menu Bar............................................................................................................ 4-6
General............................................................................................................................... 4-6
Commands ......................................................................................................................... 4-6
The Sample List Menu Bar Edit Menu .............................................................................. 4-6
The Sample List Menu Bar Samples Menu........................................................................ 4-7
Pop-up Menus ............................................................................................................................. 4-10
General............................................................................................................................. 4-10
Pop-up Menu Commands Not Available on the Sample List Menu Bar ......................... 4-11
Opening an Existing Sample List ................................................................................................ 4-11
Saving a Sample List................................................................................................................... 4-12
Printing Sample Lists .................................................................................................................. 4-13
Creating Sample Lists ................................................................................................................. 4-13
Creating a Sample List in the MassLynx Window........................................................... 4-13
Creating a Sample List by Copying a Spreadsheet .......................................................... 4-13
Importing a Worksheet into the Sample List Editor .................................................................... 4-14
General............................................................................................................................. 4-14
To Import a Worksheet .................................................................................................... 4-14
Creating Import Worksheet Files ..................................................................................... 4-15
Access .............................................................................................................................. 4-16
Excel ................................................................................................................................ 4-17
Notepad ............................................................................................................................ 4-17
Importing Data ............................................................................................................................ 4-17
General............................................................................................................................. 4-17
To Import Data................................................................................................................. 4-18
Creating Import Data Files............................................................................................... 4-18
Access .............................................................................................................................. 4-18
Excel ................................................................................................................................ 4-19
Notepad ............................................................................................................................ 4-19
Sample List Formats.................................................................................................................... 4-19
General............................................................................................................................. 4-19
Loading an Existing Sample List Format......................................................................... 4-20
Saving a Sample List Format ........................................................................................... 4-20
Changing the Sample List Format.................................................................................... 4-21
Manipulating the Sample List Editor Display ............................................................................. 4-23
Selecting Areas of the Display ......................................................................................... 4-23
Editing Data in Fields.................................................................................................................. 4-24
General............................................................................................................................. 4-24
Fields Containing Files .................................................................................................... 4-24
Fields with Multiple Options ........................................................................................... 4-25
Directly-Editable Cells..................................................................................................... 4-26
Editing Data in an Area ............................................................................................................... 4-26
Inserting Rows in a Sample List.................................................................................................. 4-26
Adding Samples to the Bottom of a Sample List ........................................................................ 4-27
Cutting Data ................................................................................................................................ 4-27
Cutting Data from a Directly-Editable Cell ..................................................................... 4-27
Cutting the Contents of Multiple Cells............................................................................. 4-27
Cutting Samples from the Sample List............................................................................. 4-27
Deleting Data............................................................................................................................... 4-28
Deleting Data from Cells.................................................................................................. 4-28
Deleting Data from Columns ........................................................................................... 4-28
Deleting Rows.................................................................................................................. 4-28
Deleting Data from the Whole Sample List ..................................................................... 4-29
Pasting Data................................................................................................................................. 4-29
Pasting Data into a Directly-Editable Cell ....................................................................... 4-29
Pasting into Multiple Cells............................................................................................... 4-29
Pasting Samples into the SLE .......................................................................................... 4-29

Page 4-2
 Sample Lists

Sorting the Rows in a Sample List .............................................................................................. 4-30

Specifying the Number of Injections for Each Sample ............................................................... 4-30
Action On Error .......................................................................................................................... 4-31
Updating the Sample List from the AutoSampler Bed Layout.................................................... 4-32
The AutoSampler Bed Layout Dialog ............................................................................. 4-32

Illustrations
Figure 4.1 Typical MassLynx Window........................................................................................ 4-5
Figure 4.2 The Sample List Menu Bar Edit Menu ....................................................................... 4-6
Figure 4.3 The Sample List Samples Menu ................................................................................. 4-7
Figure 4.4 The Samples, Fill sub-menu ....................................................................................... 4-7
Figure 4.5 The Samples, Clear sub-menu .................................................................................... 4-8
Figure 4.6 The Samples, Column sub-menu ................................................................................ 4-8
Figure 4.7 The Samples, Format sub-menu.................................................................................. 4-9
Figure 4.8 The Samples, Sort sub-menu .................................................................................... 4-10
Figure 4.9 Pop-up menu showing all possible options............................................................... 4-11
Figure 4.10 The Open File dialog .............................................................................................. 4-12
Figure 4.11 The Save As dialog ................................................................................................. 4-12
Figure 4.12 The Samples dialog................................................................................................. 4-13
Figure 4.13 The Import Worksheet dialog ................................................................................. 4-14
Figure 4.14 The Import Data dialog........................................................................................... 4-18
Figure 4.15 The Load Sample List Format dialog ..................................................................... 4-20
Figure 4.16 The Save Sample List Format dialog ...................................................................... 4-21
Figure 4.17 The Summary Information dialog........................................................................... 4-21
Figure 4.18 The Customize Field Display dialog....................................................................... 4-22
Figure 4.19 The Real Number Field Properties dialog............................................................... 4-23
Figure 4.20 The Field Properties dialog..................................................................................... 4-23
Figure 4.21 The Select File dialog ............................................................................................. 4-24
Figure 4.22 The Samples dialog................................................................................................. 4-27
Figure 4.23 The Cut or Delete dialog......................................................................................... 4-28
Figure 4.24 The Paste Option dialog.......................................................................................... 4-29
Figure 4.25 The Paste Area or Samples dialog .......................................................................... 4-29
Figure 4.26 The Number Of Injections dialog ........................................................................... 4-30
Figure 4.27 The Action On Error options .................................................................................. 4-31
Figure 4.28 Typical Autosampler Bed Layout dialog ................................................................ 4-33
Figure 4.29 The Autosampler Bed Layout dialog pop-up menu ................................................ 4-34

Page 4-3
Sample Lists

Page 4-4
 Sample Lists

Introduction
The MassLynx Sample List is a list of the samples available for analysis by the mass spectrometer,
or by sample processing. This Chapter explains how to create Sample Lists, how to change the
appearance of a Sample List, how to enter data in the Sample List and how to import Sample Lists
generated by other packages, e.g. Excel.

When MassLynx is opened, the MassLynx Window is invoked; the Sample List is displayed in the
Window; this display is, in fact, the Sample List Editor (SLE), which operates in a similar manner
to a standard spreadsheet application, e.g. Excel.

Figure 4.1 Typical MassLynx Window

The display format is customizable, allowing the User to define which fields of the Sample List
file are displayed; these configurations can be saved to named format fields for later use.

Note:
If a Waters or Gilson autosampler is installed, and controlled via MassLynx, then the Bed Layout
and Plate Layout window can be invoked. The Sample List can be updated from the plate layout,
see the “Updating the Sample List from the AutoSampler Bed Layout” section, on page 4-32, for
details.

Page 4-5
Sample Lists

The Sample List Menu Bar

General

The Sample List Menu Bar is displayed above the SLE; it contains three commands, and two
menus, associated with the Sample List.

Commands

Spectrum Invokes the Spectrum Window; the spectrum displayed will be the current
default spectrum (this will be either the last spectrum viewed, or if
acquisition is in progress, the last spectrum acquired). If the Spectrum
Window is already on display, it becomes the current Window. See
Chapter 7, “Spectrum”, for details.

Chromatogram Invokes the Chromatogram Window; see Chapter 6, “Chromatogram”, for

details. The chromatogram displayed will be the Total Ion Current (TIC)
chromatogram of the current data file (unless the Mass Chromatogram
option has been selected on the Options, System dialog, and the files
selected contain MUX data, see Chapter 3, “The MassLynx Window and
Related Information” for details). If the Chromatogram Window is
already on display it becomes the current window.

Map Creates a data file Map; see Chapter 9, “Map”, for details.

The Sample List Menu Bar Edit Menu

Figure 4.2 The Sample List Menu Bar Edit Menu

Cut Cuts selected data from the Sample List, and places it on the clipboard.
There are three types of “cut”, refer to the “Cutting Data” section, on
page 4-27, for details.

Copy Copies selected data from the Sample List, and places it on the clipboard.

Paste Pastes the contents of the clipboard into the Sample List. There are three
types of “paste”, refer to the “Pasting Data” section, on page 4-29, for
details.

Delete Deletes the contents of the selected cells.

Select All Selects the whole Sample List.

Page 4-6
 Sample Lists

The Sample List Menu Bar Samples Menu

Figure 4.3 The Sample List Samples Menu

Add Adds samples to the bottom of a Sample List, see the “Adding Samples to
the Bottom of a Sample List” section, on page 4-27.

Insert Inserts samples in the Sample List, see the “Inserting Rows in a Sample
List” section, on page 4-26.

Delete Deletes selected sample(s) from the Sample List and removes the row(s)
from the SLE. If only cells are selected, data is deleted from the cells
without removing them from the SLE.

Fill Invokes the Fill sub-menu.

Down Fills a selected Sample List range with the first element in each column,
see the “Editing Data in an Area” section, on page 4-26.

Series Fills a selected Sample List range with series data, e.g. if the first cell in a
column is bottle 1, the next will be bottle 2, then bottle 3, etc., see the
“Editing Data in an Area” section, on page 4-26.

Figure 4.4 The Samples, Fill sub-menu

Page 4-7
Sample Lists

Clear Invokes the Clear sub-menu.

Selected Clears (empties) data from the currently selected cell(s), see the “Deleting
Data from Cells” section, on page 4-28.

All Clears (empties) data from the whole Sample List, see the “Deleting Data
from the Whole Sample List” section, on page 4-29.

Figure 4.5 The Samples, Clear sub-menu

Column Invokes the Column sub-menu.

Properties Allows the name and display properties for the current column to be
changed, see the “Editing Field (Column) Properties” section, on
page 4-22.

Hide Hides the current column, see the “Removing a Column from the Sample
List Display” section, on page 4-22.

Figure 4.6 The Samples, Column sub-menu

Page 4-8
 Sample Lists

Format Invokes the Format sub-menu.

Customize Allows the User to choose the columns to be displayed in the SLE, see the
“Displaying Specific Columns” section, on page 4-21.

Load Allows an existing Sample List display format to be loaded into the
MassLynx Window, see the “Loading an Existing Sample List Format”
section, on page 4-20.

Save Allows the current Sample List display format to be saved, see the “Saving
a Sample List Format” section, on page 4-20.

Figure 4.7 The Samples, Format sub-menu

Sort Invokes the Sort sub-menu.

Sample Type Sorts the SLE rows by Sample Type.

Sample Group Sorts the SLE rows by Sample Group.

Acquisition Sorts the SLE rows by Acquisition File name.

File
Order for Orders the Sample List so that the bottle numbers are in a configuration on
MUX Wells which MUX can be used.

Page 4-9
Sample Lists

Figure 4.8 The Samples, Sort sub-menu

Number of Invokes the Samples dialog; this allows the User to specify the number of
Samples samples present in a Sample List, see the “Creating a Sample List in the
MassLynx Window” section, on page 4-13. If samples are to be removed,
a warning is given.

Number of Allows the User to specify the number of Injections to be taken of each
Injections sample, see the “Specifying the Number of Injections” section, on
page 4-30.

AutoSampler Invokes the AutoSampler Bed Layout dialog, if an appropriate

Bed Layout Autosampler has been installed, see the “Updating the Sample List from
the AutoSampler Bed Layout” section, on page 4-32.

Pop-up Menus
General

Right-clicking on a cell in the SLE invokes a pop-up menu which provides shortcuts to the most
commonly used Sample List Menu Bar commands. The invoked menu will depend on the type of
cell currently selected; commands that are not allowed are not displayed. Refer to the “The
Sample List Menu Bar” section, on page 4-6, for details of the Sample List Menu Bar commands.
Certain of the pop-up menu commands are unique to the pop-up menu and are not available on the
Sample List Menu Bar; refer to the “Pop-up Menu Commands Not Available on the Sample List
Menu Bar” section, on page 4-11, for details.

Page 4-10
 Sample Lists

Figure 4.9 Pop-up menu showing all possible options

Pop-up Menu Commands Not Available on the Sample List Menu

Bar

Browse This command is available when the selected cell can contain a file; it
invokes the Select File dialog. The User can select a file; the name of the
file will then be displayed in the cell.

Edit Launches the application to edit the particular file; e.g., selecting Edit
when an Inlet File field is selected will launch the Inlet Editor dialog.

View Launches the application to view the particular file; e.g. selecting View
when a Structure field is selected will launch the molecule viewer.

Opening an Existing Sample List

1. Select the Tool Bar button or select the Menu Bar File, Open command. This invokes
the Open file dialog, see Figure 4.10.

2. Select the required Sample List data file (of type .spl).

3. Select the Open button; the file is opened.

Page 4-11
Sample Lists

Figure 4.10 The Open File dialog

Saving a Sample List

1. Select the Tool Bar button or select the Menu Bar File, Save or File, Save As
command. If this is a newly created Sample List, or the File, Save As command has been
selected, the standard Windows Save As dialog is invoked.

Figure 4.11 The Save As dialog

2. Type the required name into the File name: box.

3. Select the Save button, the file is saved, as type .spl.

Page 4-12
 Sample Lists

Printing Sample Lists

1. Select the Tool Bar button or select the Menu Bar File, Print command; this invokes
the standard Windows Print dialog.

2. Select the required options.

3. Select the OK button.

Creating Sample Lists

Creating a Sample List in the MassLynx Window

1. Select the Tool Bar button or select the Menu Bar File, New command; a Sample List
with one default row will be created.

2. Either:

a. Select the Sample List Menu Bar Samples, Add command; the Samples dialog is
invoked.

Figure 4.12 The Samples dialog

b. Enter the required number of rows in the Number of Samples box and select the OK
button; a Sample List with the required number of rows is created.

Or:

c. Use the keyboard Insert key to add rows to the Sample List as required.

3. Change the default data in the cells as required, see the “Editing Data in Fields” section, on
page 4-24.

Creating a Sample List by Copying a Spreadsheet

Spreadsheets created in other Windows applications can be copied into the Sample List Editor.

1. Select the Tool Bar button or select the Menu Bar File, New command; a Sample List
with one default row will be created.

2. Add rows and columns to the Sample List so that it matches the number of rows and columns
in the other Windows application. If this is not done data may be lost.

Page 4-13
Sample Lists

3. Select the relevant area in the other Windows application and copy it.

4. Either:

a. In the Sample List editor, click on the cell at the top left corner of the paste area and
select the Sample List Menu Bar Edit, Paste command.

Or:

b. In the Sample List editor, right-click on the cell at the top left corner of the paste area and
select Paste from the invoked menu.

Importing a Worksheet into the Sample List Editor

General

Sample Lists can be created in a number of other packages and imported into the MassLynx
Sample List Editor. File types that can be imported are:

• OpenLynx batch files.

• Sample List files from earlier versions of MassLynx.

• Access 97 & 2000.

• Tab and Comma delimited text files.

• Excel 97& 2000, and Excel 5.0, 6.0 and 7.0 files.

To Import a Worksheet

1. Select the Menu Bar File, Import Worksheet command; the Import Worksheet dialog is
invoked.

Figure 4.13 The Import Worksheet dialog

2. Select the required file, or type a file name in the File name: box.

Page 4-14
 Sample Lists

3. The default file type is Generic Batch Files (*.OLB), created in OpenLynx; select a different
file type in the Files of type: list box, if required.

4. Select the Open button.

Note:
When a worksheet is imported, the sample list columns displayed will not change, and columns
may need to be added/removed from the display to see all the imported data.

Creating Import Worksheet Files

The following sections contain information on how to create Worksheet files suitable for
importing into MassLynx.

For all types of file, fields (columns) must have the same name as in the list below, although they
can be defined in any order. For Access 97 the data type must also match. The names correspond
to the name in brackets on the Customize Field Display dialog, see the “Displaying Specific
Columns” section, on page 4-21.

VERSION Double FILE_NAME Text

FILE_TEXT Text MS_FILE Text

MS_TUNE_FILE Text INLET_FILE Text

INLET_PRERUN Text INLET_POSTRUN Text

INLET_SWITCH Text AUTO_FILE Text

PROCESS Text PROCESS_PARAMS Text

PROCESS_OPTIONS Text PROCESS_ACTION Text

SAMPLE_LOCATION Text JOB Text

TASK Text USER Text

SUBMITTER Text CONDITIONS Text

TYPE Text IDENTIFICATION Text

CONC_A to CONC_T Text WAVELENGTH_A to Double

WAVELENGTH_J

MASS_A to MASS_T Text FRACTION_MASS Double

INJ_VOL Double STOCK_DIL Double

USER_DIVISOR_1 Double USER_FACTOR_1 Double

USER_FACTOR_2 Double USER_FACTOR_3 Double

SPARE_1 to SPARE_5 Text HPLC_FILE Text

Index Double ACQU_PROCESS Text

Page 4-15
Sample Lists

ACQU_PROCESS_PARAMS Text ACQU_PROCESS_OPTIONS Text

SAMPLE_GROUP Text FRACTION_FILE Text

FRACTION_1 to FRACTION_4 Text QUAN_REFERENCE Text

Access

General

When the table is created it must be called ANALYSIS.

It is recommended that the design view is used when creating a new table, this allows the User to
define the field data type.

The first column must be called Index as the primary key. The Index column defines the order in
which the rows will be displayed in MassLynx. The AutoNumber data type could be used to set up
this column in Access.

Column headings must match those above. Other columns may be present but they will not be
imported into the Sample List.

To Define the Data Type as a Double

1. Select Number from the list box in the Data Type column.

2. On the general page, at the bottom left of the window, click on Field Size and select Double
from the drop-down list box.

To Save in Access Format

The table can be saved as an access database by selecting the Menu Bar File, Save command, and
can be imported into MassLynx in this format. Tables can also be saved as tab or comma
delimited files for importing into MassLynx; see the “To Save in Tab or Comma Delimited
Format” section, below.

To Save in Tab or Comma Delimited Format

For Access 97:

1. Select the Menu Bar File Save As/Export command, select the To an External File or
Database option and select the OK button.

2. Select the required directory from the browser displayed, select Text files
(*.txt;*.csv;*.tab;*.asc) from the Save as type list box and then select the Export button.

3. Make sure the Delimited option is selected and select the Next button.

4. Select the Include Field Names on First Row option, select the type of delimiter to use and
select the Next button.

5. Enter the required file name.

6. Select the Finish button.

Page 4-16
 Sample Lists

For Access 2000:

1. With the ANALYSIS table open, select the Menu Bar File, Export command.

2. Select the required directory from the browser displayed, select Text files
(*.txt;*.csv;*.tab;*.asc) from the Save as type list box and then select the Save button.

3. Make sure the Delimited option is selected and select the Next button.

4. Select the Include Field Names on First Row option, select the type of delimiter to use and
select the Next button.

5. Enter the required file name.

6. Select the Finish button.

If files are saved as comma or tab delimited, they must be imported into MassLynx as comma or
tab delimited files.

Excel

The first column must be called Index, the other column headings must match those shown on
page 4-14.

Select the area containing the data to be imported, including the column headings, and name the
area ANALYSIS. To do this, select the Insert, Name, Define command, type ANALYSIS and
select OK.

Leave all cells in General format.

For a text field containing only numeric data an apostrophe ( ‘ ) must be inserted in front of the
number.

If the file is to be saved as tab or comma delimited, Excel will only allow one sheet to be saved. If
the current workbook contains more than one worksheet, each worksheet must be saved as a
separate text file.

Notepad

The first column must be called Index, the other column headings must match those shown on
page 4-14.

Type in the field name/value and then a comma (or press tab for tab delimited files) and enter the
next value. End each line with a carriage return.

Text fields should be enclosed in quotes.

Importing Data
General

Sample List data can be created in a number of other packages and imported into MassLynx. The
file types supported are:

• ACCESS 97 & 2000.

Page 4-17
Sample Lists

• Tab and Comma delimited text files.

• Excel 97 & 2000 and Excel 5.0, 6.0 and 7.0 files.

To Import Data

1. In MassLynx, ensure that the correct number of rows and columns required to accept the data
is displayed. Data will be lost if this is not done.

2. Select the MassLynx Menu Bar File, Import Data command; the Import Data dialog is
invoked.

Figure 4.14 The Import Data dialog

3. Select the required file, or type a file name in the File name: box

4. The default file type is Excel 5.0 (*.XLS); select a different file type in the Files of type: list
box, if required.

5. Select the Open button.

Creating Import Data Files

The following sections contain information on how to create files suitable for importing into
MassLynx.

For all types of file:

• Fields must not have column headings.

• Fields must be in the same order as they are to appear in the MassLynx Sample List.

Access

When the table is created it must be called ANALYSIS.

It is recommended that the design view is used when creating a new table, this allows the field data
type to be defined. The field names do not have to correspond to the field names in MassLynx
(but the order must be the same).

Page 4-18
 Sample Lists

To define the data type as a double, see the “To Define the Data Type as a Double” section, on
page 4-16.

To save in tab or comma delimited format, follow the instructions in the “To Save in Tab or
Comma Delimited Format” section (on page 4-16), except for step 4, where the Include Field
Names on First Row option must not be selected.

If files are saved as comma or tab delimited, they must be imported into MassLynx as comma or
tab delimited files.

Excel

Select the area containing the data to be imported, including the column headings, and name the
area ANALYSIS. To do this, select the Menu Bar Insert, Name, Define command, type
ANALYSIS and select the OK button. The column headings do not have to correspond to the
field names in MassLynx, but the order must be the same.

Leave all cells in General format.

For a text field containing only numeric data an apostrophe ( ‘ ) must be inserted in front of the
number.

If the file is to be saved as tab or comma delimited, Excel will only allow one sheet to be saved. If
the current workbook contains more than one worksheet, each worksheet must be saved as a
separate text file. Column headings or blank rows should NOT be included in the ANALYSIS
area when saving to a text file.

Notepad

Type in the field name/value, then a comma (or press the tab key for tab delimited files) and enter
the next value. End each line with a carriage return. Column headings should not be included, but
the fields must be in the same order as in MassLynx.

Text fields should be enclosed in quotes.

Sample List Formats

General

Sample List format details are stored in Sample List format files (of type .fmt). MassLynx is
supplied with some default formats. Sample List formats can be edited (see the “Changing the
Sample List Format” section, on page 4-21), and then saved and retrieved as required.

Note.
If a Sample List is open, and a new format is loaded, this will replace the current format.

Page 4-19
Sample Lists

Loading an Existing Sample List Format

1. Select the Sample List Menu Bar Samples, Format, Load command. This invokes the Load
Sample List Format dialog.

Figure 4.15 The Load Sample List Format dialog

2. Select the required format in the list box. If the format is not present in the current directory
(displayed above the list box), select the Browse button and locate the file from the invoked
dialog.

Note:
1. The name displayed in the list box is the title entered in the Summary Information
dialog Title: text box when the Sample List format was saved; this is not necessarily
the file name, see the “Saving a Sample List Format” section, below.

2. Any Description displayed is that entered in the Summary Information dialog

Description: text box when the Sample List format was saved.

3. Select the OK button.

Saving a Sample List Format

1. Format the spreadsheet to include the required fields; see the “Changing the Sample List
Format” section, on page 4-21.

2. Select the Sample List Menu Bar Samples, Format, Save command; this invokes the Save
Sample List Format dialog.

Page 4-20
 Sample Lists

Figure 4.16 The Save Sample List Format dialog

3. Enter the file name, select a location and select the Save button. The Summary Information
dialog is invoked.

Figure 4.17 The Summary Information dialog

4. Enter details as required In the Title: and Description: text boxes, and select the OK button.
Title: is the text that will appear in the Load Sample List Format dialog list box when
loading formats. Description: is the text that will appear in the Load Sample List Format
dialog Description box.

Changing the Sample List Format

Changing the Column Width

Column widths can be changed, in the same way as for any Windows spreadsheet. Position the
mouse pointer on the line between two column headings until a double headed arrow appears, then
click and drag until the column is the required width.

Displaying Specific Columns

Many different columns of information can be displayed in the Sample List Editor; the User can
select which columns are currently displayed.

1. Either:

a. Select the Sample List Menu Bar Samples, Format, Customize command

Or:

Page 4-21
Sample Lists

b. Right-click on the Sample List and select the Customize Display command from the
invoked menu.

In either case, the Customize Field Display dialog is invoked.

Figure 4.18 The Customize Field Display dialog

2. Select the box next to a field to display the field in the Sample List.

3. To move the position of a field, select the field in the list and select the Move or
buttons until the field is in the required position.

Removing a Column from the Sample List Display

1. Select the column to be removed.

2. Select the Sample List Menu Bar Samples, Column, Hide command. The column is
removed from the SLE, however its contents are not removed, and data is still automatically
propagated when samples are added or inserted.

Editing Field (Column) Properties

1. Either:

a. Select the column heading and select the Sample List Menu Bar Samples, Column,
Properties command.

Or:

b. Right-click on the column heading and select the Properties command from the invoked
menu.

In either case, a Field Properties dialog is invoked.

Note:
There are two types of Field Properties dialog; one is for real number fields and allows the
number of decimal places displayed to be to be specified (see Figure 4.19), the other dialog is for
all other fields, see Figure 4.20.

Page 4-22
 Sample Lists

Figure 4.19 The Real Number Field Properties dialog

Figure 4.20 The Field Properties dialog

2. To change the column name, type a new name into the Field name: box.

3. To change the alignment of text in the column, choose Left, Right or Center from the
Alignment: list.

4. For real number fields only, enter the number of decimal places to be displayed (maximum
four) in the Decimal Places box; alternatively, scroll to the required number using the arrows,
, see Figure 4.19.

Manipulating the Sample List Editor Display

Selecting Areas of the Display

General

Display areas may be selected with the mouse, the keyboard or a combination of both of these
methods.

With the Mouse

To select: Click on:

A single cell. The required cell.

A block of cells. The first cell in the block; hold down the mouse button and drag
until the required cells are highlighted.

A row. The row number.

A column. The column heading.

The whole Sample List. The box at the top left corner of the Sample List.

Page 4-23
Sample Lists

With the Keyboard

Position the cursor at the top left corner of the area to be selected, hold down the shift key and use
the arrow keys to select an area.

Editing Data in Fields

General

Data can be changed in various ways, the following sections describe the most common methods.

Fields Containing Files

For the following field types, files created and saved in the current project can be included in a cell
by double-clicking on it; this invokes the Select File dialog. Select the required file in the normal
Windows manner.

Figure 4.21 The Select File dialog

Field: File Type: Directory:

MS File *.exp (*.mdb for TDAT instruments) Acqudb

Parameter file *.olp (Default; this depends on the current specified process, MassLynx
if the process is known to MassLynx a different file
extension may be presented.)

Page 4-24
 Sample Lists

Field: File Type: Directory:

Inlet File *.wat (Waters 2690), *.w60 (Waters 600), Acqudb

*.w27 (Waters 2700), *.w29 (Waters 2790),
*.clc (Waters Cap LC), *.gil (Gilson), *.h11 (HP 1100),
*.h50(HP 1050), *.h68 (HP 6890), *.h90 (HP 1090),
*.szu (Shimadzu), *.jas (Jasco 900), *j15 (Jasco 1500),
*.asx (Cetac ASX500), *.as1 (Cetac ASX100),
*.ct2 (CTC A200S)

Note:
A custom “browse” dialog is displayed that only contains
files valid for the current configuration.

MS Tune File *.ipr (*.dbf for TDAT instruments) Acqudb

Inlet Prerun See Inlet File. Acqudb

Inlet Postrun See Inlet File. Acqudb

Inlet Switch See Inlet File. Acqudb

Autofile See Inlet File. Acqudb

Acqu Parameter *.* Acqudb

Fraction File *.frc Acqudb

HPLC File *.lca Acqudb

Structure *.mol MassLynx

Fields with Multiple Options

For the following field types, select one of the displayed options from the list box invoked by
double-clicking on a cell in the field:

Field: Options:

Process This field contains many options. Many of the available options are not
valid Sample List processes; care should be taken to select an option that is
valid for the current application.

Control Analyte, Control.

Sample Type Analyte, Blank, QC, Standard.

Action on Error Ignore Error, Suspend this Batch, Suspend All batches, Delete this Batch.

Fraction Trigger Mass A to T, Wavelength A to J, Mass TIC, PDA TIC, Analog 1 to 4,

UV 1 to 4, No Trigger.

Page 4-25
Sample Lists

Directly-Editable Cells

For fields other than those described above, to edit data in a cell, double-click on the cell and type
in a new value. When editing data in a single cell, cut, copy, paste, etc. can be performed in the
usual manner, or a single right-mouse click will invoke a menu with a range of commands.

Note:
Selecting a cell and pressing the F2 key allows virtually any cell to be edited directly without
invoking a dialog.

Editing Data in an Area

Selecting a Sample List Editor area more than one row in depth and selecting the Sample List
Menu Bar Samples, Fill, Down command, will copy the contents of the first selected cell(s) and
paste them into the cells below.

Selecting an area more than one row in depth and selecting the Sample List Menu Bar Samples,
Fill, Series command, will fill the selected range with series data, e.g. if the first cell in a column
is bottle1, the next will be bottle2, bottle3, etc. If there is more than one number in a field then
only the last number is incremented when Samples, Fill, Series is selected, e.g. for Sample1run1,
when Samples, Fill, Series is selected, the next field will be Sample1run2, etc.

Inserting Rows in a Sample List

To insert a single row, position the cursor on the row above which the new row is to be inserted.

Either:

a. Select the Sample List Menu Bar Samples, Insert command.

Or:

b. Press the Insert keyboard key.

Or:

c. Right-click and select Insert from the invoked menu.

If more than one row is selected the same number of rows will be inserted above the first row of
the selection. If a column has been selected, the same number of rows as were originally in the
column is inserted before the first row. The data inserted into these new rows will continue the
series from the row above the selection.

For example, in the following example, selecting the two rows highlighted in the first picture and
inserting using any of the above commands, will give the second picture.

Page 4-26
 Sample Lists

Adding Samples to the Bottom of a Sample List

To add rows to the bottom of a Sample List:

1. Select the Sample List Menu Bar Samples, Add command; the Samples dialog is invoked.

Figure 4.22 The Samples dialog

2. Enter the number of samples to be added to the Sample List in the Number of Samples box;
the number can be typed, or incremented using the arrows, .

3. Select the OK button; the required number of samples, with default data from the row above,
is added to the Sample List.

Cutting Data
Cutting Data from a Directly-Editable Cell

Double-click on a cell; the cell now becomes an edit box; part, or all, of the cell’s contents can
then selected and cut using the Sample List Menu Bar Edit, Cut command.

Cutting the Contents of Multiple Cells

Any area of cells may be selected, and the contents cut using the Sample List Menu Bar Edit, Cut
command.

Cutting Samples from the Sample List

To cut samples from the Sample List, select the row(s) to be cut and select the Sample List Menu
Bar Edit, Cut command; the Cut or Delete dialog is invoked.

Page 4-27
Sample Lists

Figure 4.23 The Cut or Delete dialog

Data or Samples
Frame
Data in Cuts data from the selected row(s) without removing the row(s) from the
Selected Range SLE.

Samples from Removes the selected row(s) from the SLE.

Sample List
Note:
All the rows cannot be cut, or deleted from the SLE; there must always be at least one row in the
Sample List.

Deleting Data
Deleting Data from Cells

1. Select the cell(s) from which data is to be deleted.

2. Either:

a. Select the Sample List Menu Bar Samples, Delete command.

Or:

b. Select the Sample List Menu Bar Samples, Clear, Selected command.

Deleting Data from Columns

1. Select the column(s) from which data is to be deleted.

2. Either:

a. Select the Sample List Menu Bar Samples, Delete command.

Or:

b. Select the Sample List Menu Bar Samples, Clear, Selected command.

Deleting Rows

1. Select the row(s), to be deleted.

2. Select the Sample List Menu Bar Samples, Delete command.

Page 4-28
 Sample Lists

Deleting Data from the Whole Sample List

Select the Sample List Menu Bar Samples, Clear, All command.

Pasting Data
Pasting Data into a Directly-Editable Cell

Double-click on the cell; the cell now becomes an edit box; paste into the cell using the Sample
List Menu Bar Edit, Paste command.

Note:
Pressing the F2 key allows instant editing in most types of cell.

Pasting into Multiple Cells

Previously cut multiple cells (see the “Cutting the Contents of Multiple Cells” section, on
page 4-27) can be pasted into a selected area of the SLE using the Sample List Menu Bar Edit,
Paste command. If the selected area differs from the cut area, the Paste Option dialog is invoked.

Figure 4.24 The Paste Option dialog

If the Yes button is selected, any data that cannot be accommodated in the selected area will not be
pasted.

Pasting Samples into the SLE

Previously cut Sample rows (see the “Cutting Samples from the Sample List” section, on
page 4-27) can be pasted into the SLE using the Sample List Menu Bar Edit, Paste command; the
Paste Area or Samples dialog is invoked.

Figure 4.25 The Paste Area or Samples dialog

Page 4-29
Sample Lists

Paste Data Frame

In selected Pastes the Sample row data into the currently-selected range of cells. The
Range Paste Option dialog may be invoked when using this option, see the
“Pasting into Multiple Cells” section, on page 4-29.

Paste New
Samples Frame
After Selected Inserts the complete Sample row(s) into the SLE immediately after the row
Range containing the currently-selected cell(s).

Before Selected Inserts the complete Sample row(s) into the SLE immediately before the
Range row containing the currently-selected cell(s).

Sorting the Rows in a Sample List

The order of the rows in a Sample List can be sorted as follows:

To order by Sample Type, select the Sample List Menu Bar Samples, Sort, Sample Type
command.

To order by Sample Group, select the Sample List Menu Bar Samples, Sort, Sample Group
command.

To order by Acquisition File, select the Sample List Menu Bar Samples, Sort, Acquisition File
command.

To order so that the bottle numbers are in a configuration on which MUX can be used, select the
Sample List Menu Bar Samples, Sort, Order for MUX Wells command.

Specifying the Number of Injections for Each

Sample
Select the Sample List Menu Bar Samples, Number of Injections command; the Number Of
Injections dialog is invoked.

Figure 4.26 The Number Of Injections dialog

Enter the required number of injections to be taken of each Sample and select the OK button.

This controls the number of injections taken from one bottle; for example, for the following
Sample List:

Page 4-30
 Sample Lists

If Number of Injections is changed to 2, the Bottle column selected, and the Sample List Menu
Bar Samples, Fill, Series command selected, the Sample List changes to:

This indicates that injections 1 and 2 are taken from bottle 1, injections 2 and 3 are taken from
bottle 2, etc.

If the User subsequently changes the number of Samples (using the Sample List Menu Bar
Samples, Number of Samples command), a Sample is added N times (where N is the number of
injections) for each bottle; e.g. if N=5, five bottles will be added for each new Sample. Up to ten
injections can be specified.

Action On Error
The Action On Error field allows the User to define what happens to a batch if an error occurs.
Select the Customize Field Display dialog Action On Error option (see the “Displaying Specific
Columns” section, on page 4-21) to display the column on the Sample List. Double-clicking on a
cell invokes a list box ; select one of the options.

Figure 4.27 The Action On Error options

Ignore Error Ignore the error and continue with the acquisition.

Suspend This Pauses the current batch and continues with the next batch in the
Batch MassLynx Queue.

Suspend All Pauses all batches.

Batches
Delete This Deletes the current batch from the queue and continues with the next one.
Batch
If no action is chosen, Ignore Error is selected by default.

Page 4-31
Sample Lists

Updating the Sample List from the AutoSampler

Bed Layout
The AutoSampler Bed Layout Dialog

Description

If any of the following autosamplers is installed, and controlled via MassLynx, the AutoSampler
Bed Layout dialog may be invoked by selecting the Sample List Menu Bar Samples,
AutoSampler Bed Layout command:

• Gilson 215

• Gilson 231XL

• Gilson 232XL

• Gilson 233XL

• Waters 2700

• Waters 2795HT

• Waters 2747

• Waters 2767

Note:
1. Autosamplers are selected using the Inlet Method dialog, refer to the “MassLynx NT Guide
to Inlet Control” for details.

2. The appearance of the Autosampler Bed Layout dialog depends on the current autosampler,
however the controls described in this section are common to all types.

The Autosampler Bed Layout dialog allows sample locations to be entered in the Sample List.

Page 4-32
 Sample Lists

Bed Layout

Currently-selected plate

Plate Control

Figure 4.28 Typical Autosampler Bed Layout dialog

Inserts the samples, currently selected in the Tray control, in the SLE.

Replaces the samples currently selected in the SLE, with those currently-
selected in the Tray control.

Adds the samples currently selected in the Tray control, to the SLE.

Bed Layout Provides a visual description of the current autosampler bed layout. Click
on a plate to make it the current plate, see Currently-selected plate.

Currently- Displays the plate currently-selected in the Autosampler Bed Layout. In

selected plate Figure 4.28, the third plate on the second row is current.

Selects all the vials on the current plate.

Deselects all the vials on the current plate.

Plate Control Provides a visual representation of the currently-selected plate. Clicking

on a vial position selects/deselects the vial. When deselected, the vial is
colored black, ; when selected, the vial is colored green, .

Page 4-33
Sample Lists

The Autosampler Bed Layout Dialog Pop-Up Menu

Right-clicking on the Autosampler Bed Layout dialog invokes the following pop-up menu.

Figure 4.29 The Autosampler Bed Layout dialog pop-up menu

The commands on this menu have the same functions as the Autosampler Bed Layout dialog
buttons:

Button Menu Command Function

Insert Inserts the samples, currently selected in the Tray control, in
the SLE.

Add Adds the samples currently selected in the Tray control, to the
SLE.

Replace Replaces the samples currently selected in the SLE, with those
currently-selected in the Tray control.

Select all Vials Selects all the vials on the current plate.

Un-select All Deselects all the vials on the current plate.

Vials
To Select the Bed Layout

For the Waters 2790 autosampler there is only one Bed Layout, which will be displayed
automatically.

For the Waters 2700 and Gilson autosamplers, the Bed Layout selected on the Inlet Method
dialog, Sampler Configuration page is displayed. See the relevant Instrument User’s Guide for
details.

To Select the Plate

Select a plate in the Bed Layout area; the plate number will be updated and, if the plate layout is
different from the previous one, the picture will be updated.

To Select Vials

Selected vials are shown in green, , deselected ones in black, .

To select a vial, click on a black vial. To select all vials on the plate, select the button, or
right-click on the plate and select the Select all Vials command from the invoked menu.

To Deselect Vials

To deselect a vial, click on a green vial. To deselect all vials on the plate, select the button,
or right-click on the plate and select the Un-select all Vials command from the invoked menu.

Page 4-34
 Sample Lists

To Append Samples to the Sample List

Select the required vials in the Autosampler Bed Layout dialog and select the button, or
right-click on the dialog and select the Add command from the invoked menu.

The vials selected on every plate (including the plates that are not current) will be appended to the
Sample List. The fields will be filled as though a Sample List Menu Bar Samples, Fill, Series
command has been performed, see the “Editing Data in an Area” section, on page 4-26. For
example, if the last row in the Sample List was

and the following vials from Plate 1,1 added,

the Sample List will be updated as follows:

To Insert Samples into the Sample List

Select the required vials in the Autosampler Bed Layout dialog and, in the SLE, click on the row
above which the samples are to be inserted. Select the Autosampler Bed Layout dialog
button, or right-click on the plate and select Insert from the invoked menu.

The vials selected on every plate (including the plates that are not current) will be inserted into the
Sample List. The fields will be filled as though a Sample List Menu Bar Samples, Fill, Series
command has been performed, see the “Editing Data in an Area” section, on page 4-26. For
example, if the following row is selected in the Sample List,

and the following vials from Plate 1,1 inserted,

Page 4-35
Sample Lists

the Sample List will be updated as follows:

Note:
File names may need to be updated as this operation may cause names to be duplicated.

To Replace Samples in the Sample List

Select the required vials in the Autosampler Bed Layout dialog and, in the SLE, the rows to be
replaced. The rows to be replaced must be next to each other and must match the number of
samples selected. Select the button, or right-click on the plate and select the Replace
command from the invoked menu.

The SAMPLE_LOCATION field for the selected samples will be replaced by location of the vials
selected from the plate(s). For example, if the following rows are selected in the Sample List

and replaced with the following vials from Plate 1,1,

the Sample List will be updated as follows:

Page 4-36
 The MassLynx Queue

Chapter 5 The M assLynx Queue

Page 5-1
The MassLynx Queue

Contents
Introduction ................................................................................................................................... 5-3
Starting Data Acquisition .............................................................................................................. 5-3
The Queue Bar .............................................................................................................................. 5-5
General............................................................................................................................... 5-5
Changing the Job Properties............................................................................................... 5-6
Changing the Queue Properties.......................................................................................... 5-7

Illustrations
Figure 5.1 The Start Sample List Run dialog ............................................................................... 5-3
Figure 5.2 Typical Queue Bar ...................................................................................................... 5-5
Figure 5.3 Typical Process dialog ................................................................................................ 5-6
Figure 5.4 The Queue Properties dialog ....................................................................................... 5-7

Page 5-2
 The MassLynx Queue

Introduction
When data acquisition is started, the job is submitted to the MassLynx Queue; multiple jobs may
be submitted. The Queue Bar is used to manage the jobs currently in the Queue and allows the
User to prioritize jobs, define when they are to run, etc.

Starting Data Acquisition

To start data acquisition, select the Menu Bar Run, Start command, or select the Tool Bar
button; the Start Sample List Run dialog is invoked.

Note:
If changes have been made to the Sample List, but have not been saved, a “Save changes?”
prompt is initially invoked.

When the required options have been selected, select the OK button, the job is then submitted to
the MassLynx Queue.

Figure 5.1 The Start Sample List Run dialog

Project Frame The name of the current project appears here. To acquire to a different
project, exit this dialog, open another project and start acquisition again.

Acquire Sample Acquires data for all the Samples in the List.
Data

Page 5-3
The MassLynx Queue

Auto Process Processes acquired data as specified in the Sample List Process column,
Samples see Chapter 4, “Sample Lists” for further details. This may be existing
data, or data newly acquired when the Acquire Sample Data option is
selected.

Auto Quantify Quantifies the acquired data using the method specified in the Quantify
Samples Samples dialog, see Chapter 10, “Quantify” for details. The current
method will be used if a method is not defined in the Quantify Samples
dialog. If this option is selected, the Quantify Samples dialog will be
invoked when the Start Sample List Run dialog OK button is selected.

Note:
The above three actions can be run together or independently; i.e. the User can acquire, process
and quantify data in one go, or acquire data in one run and process or quantify it at a later date.

Run Frame

From Sample Insert the sample number (from the Sample List) for the first sample to be
acquired.

To Sample Insert the sample number for the last sample to be acquired.

Priority Marks this job as a Priority process; it will be placed at the top of the
Queue, to run after the currently running job, see the “Changing the Job
Properties” section, on page 5-6.

Night Time Marks this entry this entry as a night time process, see the “Changing the
Process Queue Properties” section, on page 5-7. This option is useful for time-
consuming acquisitions that would interrupt work on smaller acquisitions
during the day.

Process Frame

Pre-Run Causes the external executable process specified in the adjacent text box to
be run and perform pre-processing on the Samples.

Post-Run Causes the external executable process specified in the adjacent text box to
be run and perform post-processing on the Samples.

Note:
Any .exe file can be run, hence this allows Users to write their own applications to perform a task
before, or after the batch is executed.

Page 5-4
 The MassLynx Queue

The Queue Bar

General

The Queue Bar is displayed in the MassLynx Window, it is invoked by selecting the Menu Bar

View, MassLynx Bar, Queue command, or the Tool Bar button. This Bar is used to
manage the MassLynx Queue.

Figure 5.2 Typical Queue Bar

The icons displayed in the Queue Bar indicate the jobs currently in the Queue.

Denotes the job currently running.

Denotes a job waiting in the queue.

Denotes a job that has been paused, see the “Changing the Job Properties”
section, on page 5-6.

Denotes a job defined as a priority process, see the “Changing the Job
Properties” section, on page 5-6.

Note:
The Queue Properties dialog Pre-emptive Scheduling option must also
be selected, see the “Changing the Queue Properties” section, on
page 5-7.

Denotes a job defined as a Night Time Process, see the “Changing the
Queue Properties” section, on page 5-7.

Note:
The Queue Properties dialog Night Time Scheduling option must also
be selected, see the “Changing the Queue Properties” section, on
page 5-7.

Page 5-5
The MassLynx Queue

Changing the Job Properties

Clicking on a job icon in the Queue Bar invokes the Process dialog for that job.

Figure 5.3 Typical Process dialog

The frame at the top of the dialog displays information about the job.

Pause Process Pauses the job; the job will not be run until this option is deselected.

Priority Process Marks this job as a Priority process; it will be placed at the top of the
Queue.

If this option is selected and a non-priority process is acquiring, the current

sample will be acquired; the current process will then be paused and the
priority process will start acquisition. When the priority process has
finished acquisition, the previous process will continue. If more than one
job is specified as a Priority Process, the jobs will be processed
chronologically.

Note:
The Queue Properties dialog Pre-emptive Scheduling option must also
be selected, see the “Changing the Queue Properties” section, on
page 5-7.

Night Time Marks this entry this entry as a night time process, see the “Changing the
Process Queue Properties” section, on page 5-7.

Note:
The Queue Properties dialog Night Time Scheduling option must also
be selected, see the “Changing the Queue Properties” section, on
page 5-7.

Select this icon to delete the process from the Queue.

Note:
The Queue is paused if the process is running when deleted.

Page 5-6
 The MassLynx Queue

Changing the Queue Properties

Select the Properties tab, at the left-hand-side of the Queue Bar, to change the Queue properties;
this invokes the Queue Properties dialog.

Figure 5.4 The Queue Properties dialog

Scheduling
Frame
Pre-emptive Defines the currently selected MassLynx Queue entry as a priority process.
Scheduling
Night Time Defines the currently selected MassLynx Queue entry as a night-time
Scheduling process. As such, the process will wait in the queue until the Night Start
Time before acquiring data.

Night Time
Settings Frame
Night Start
The start time for a night-time process, see above. Pressing the buttons
Time
will increase or decrease the time by one hour. To change the minutes
setting, click on the minutes part of the display; pressing the buttons
will now increase or decrease the time by one minute.

Night End
The end time for a night-time process, see above. Pressing the buttons
Time
will increase or decrease the time by one hour. To change the minutes
setting, click on the minutes part of the display; pressing the buttons
will now increase or decrease the time by one minute.

Restore queue on Normally, when MassLynx is closed, any processes on the MassLynx
start-up Queue are lost. Select this option to save the details of the Queue and to
restore them when MassLynx is restarted.

Page 5-7
The MassLynx Queue

Page 5-8
 Chromatogram

Chapter 6 Chromatogram

Page 6-1
Chromatogram

Contents
Getting Started............................................................................................................................... 6-7
To Display the Total Ion Current (TIC) Chromatogram .................................................... 6-7
To Display a Summed Mass Chromatogram Around a Peak in a Spectrum...................... 6-7
The Chromatogram Window......................................................................................................... 6-7
The Chromatogram Menu Bar....................................................................................................... 6-8
The Chromatogram File Menu........................................................................................... 6-8
The Chromatogram Edit Menu .......................................................................................... 6-8
The Chromatogram Display Menu..................................................................................... 6-9
The Chromatogram Process Menu ................................................................................... 6-12
The Chromatogram Window Menu ................................................................................. 6-13
The Chromatogram Tools Menu ...................................................................................... 6-14
The Chromatogram Help Menu ....................................................................................... 6-14
The Chromatogram Tool Bar ...................................................................................................... 6-15
General............................................................................................................................. 6-15
Customizing the Chromatogram Tool Bar ....................................................................... 6-16
Displaying Chromatograms......................................................................................................... 6-19
Adding or Replacing Chromatogram Traces.................................................................... 6-19
The New Chromatogram Dialog ...................................................................................... 6-20
The Mass Chromatogram dialog ...................................................................................... 6-20
TIC and BPI Chromatograms........................................................................................... 6-24
Analog Chromatograms ................................................................................................... 6-25
Aligning Analog Chromatograms .................................................................................... 6-26
The Chromatogram Pointer ......................................................................................................... 6-27
Manipulating the Display ............................................................................................................ 6-27
Altering the Range of the Horizontal Axis....................................................................... 6-27
Centering the Display Around a Particular Point ............................................................. 6-28
Altering the Range of the Intensity Axis.......................................................................... 6-29
Altering the Range of Both Axes ..................................................................................... 6-29
Setting Magnified Ranges ................................................................................................ 6-29
Deleting Magnification Ranges........................................................................................ 6-30
Restoring the Display....................................................................................................... 6-31
Setting the Display Range Defaults ................................................................................. 6-31
Controlling the Appearance of the Display ................................................................................. 6-32
General............................................................................................................................. 6-32
To Change the Display Parameters .................................................................................. 6-32
Controlling the Appearance of Peak Labels ................................................................................ 6-34
General............................................................................................................................. 6-34
To Change the Peak Annotation Parameters .................................................................... 6-34
Removing Chromatograms from the Display.............................................................................. 6-36
To Remove a Single Chromatogram Trace from the Display .......................................... 6-36
To Remove Multiple Chromatogram Traces from the Display........................................ 6-36
Real-Time Display of Chromatograms........................................................................................ 6-36
Changing the Order of Displayed Chromatograms ..................................................................... 6-37
Adding Text to the Chromatogram Display ................................................................................ 6-37
Processing Chromatograms ......................................................................................................... 6-38
General............................................................................................................................. 6-38
Processing Multiple Chromatograms ............................................................................... 6-38
Background Subtract ................................................................................................................... 6-39
General............................................................................................................................. 6-39
Performing a Background Subtract .................................................................................. 6-39
Example of Background Subtract .................................................................................... 6-40
Checking the Results of Background Subtract................................................................. 6-41
Smoothing Chromatograms......................................................................................................... 6-41
General............................................................................................................................. 6-41
The Smooth Chromatogram Dialog ................................................................................. 6-41
To Smooth a Chromatogram ............................................................................................ 6-42
Integrating Chromatograms......................................................................................................... 6-43

Page 6-2
 Chromatogram

General............................................................................................................................. 6-43
To Integrate a Chromatogram .......................................................................................... 6-44
Standard Peak Detection Parameters ............................................................................... 6-47
ApexTrack Peak Detection Parameters............................................................................ 6-52
Peak Thresholding ........................................................................................................... 6-53
To Display Information about an Integrated Peak ........................................................... 6-54
Editing Integrated Peaks .................................................................................................. 6-54
Peak Purity .................................................................................................................................. 6-57
General............................................................................................................................. 6-57
To Calculate the Peak Purity Index for a Total Ion Chromatogram................................. 6-59
Signal to Noise Ratio .................................................................................................................. 6-59
General............................................................................................................................. 6-59
The Signal To Noise dialog ............................................................................................. 6-60
To Calculate the Signal to Noise Value for a Mass Chromatogram ................................ 6-61
Combine Spectra ......................................................................................................................... 6-61
ElectroSpray Data Processing – Components ............................................................................. 6-62
General............................................................................................................................. 6-62
Component Identification ................................................................................................ 6-62
The Auto Find Components Dialog ................................................................................. 6-63
Automatic Component Finding........................................................................................ 6-64
The Component Worklist Dialog..................................................................................... 6-66
Peak Lists .................................................................................................................................... 6-69
General............................................................................................................................. 6-69
The Edit Peak List Dialog................................................................................................ 6-69
To Create a New Peak List File ....................................................................................... 6-71
To Open an Existing a Peak List File .............................................................................. 6-72
To Append Peaks to the Current Peak List ...................................................................... 6-72
To Delete Peaks from the Current Peak List.................................................................... 6-72
Reading a Peak List into a Chromatogram ...................................................................... 6-72
Chromatogram Display When Switching Between MS and MS/MS Modes.............................. 6-74
Copying To and From the Windows Clipboard .......................................................................... 6-75
General............................................................................................................................. 6-75
To Copy a Chromatogram as a Picture to the Clipboard ................................................. 6-75
To Copy a Chromatogram as a Text List to the Clipboard .............................................. 6-75
To Copy Integrated Chromatogram Peaks as a Text List to the Clipboard...................... 6-75
To Paste Information from the Windows Clipboard into a Chromatogram Window ...... 6-75
Removing Pasted Input from the Display ........................................................................ 6-76
Retention Index ........................................................................................................................... 6-76
General............................................................................................................................. 6-76
The Retention Index Table dialog.................................................................................... 6-76
To Set Up the Retention Index Table............................................................................... 6-77
To Delete the Retention Index Table ............................................................................... 6-77
To Make a Retention Index Calibration........................................................................... 6-77
To Display Retention Index Values on a Chromatogram ................................................ 6-78
To Check Retention Index Calibration Status.................................................................. 6-79

Illustrations
Figure 6.1 The Chromatogram Window ...................................................................................... 6-7
Figure 6.2 The Chromatogram File Menu.................................................................................... 6-8
Figure 6.3 The Chromatogram Edit Menu ................................................................................... 6-8
Figure 6.4 The Chromatogram Display Menu ............................................................................. 6-9
Figure 6.5 The Chromatogram Display, Range Sub-menu ........................................................ 6-11
Figure 6.6 The Chromatogram Display, Range, Center sub-menu ............................................ 6-12
Figure 6.7 The Chromatogram Process Menu............................................................................ 6-12
Figure 6.8 The Process, Components sub-menu ........................................................................ 6-13
Figure 6.9 The Chromatogram Window Menu .......................................................................... 6-13
Figure 6.10 The Chromatogram Tools Menu............................................................................. 6-14

Page 6-3
Chromatogram

Figure 6.11 Typical Retention Index Status dialog .................................................................... 6-14

Figure 6.12 The Customize Toolbar dialog................................................................................ 6-17
Figure 6.13 The New Chromatogram dialog.............................................................................. 6-20
Figure 6.14 The Mass Chromatogram dialog for full scan data ................................................. 6-21
Figure 6.15 The Mass Chromatogram dialog for Selected Ion Recording (SIR) data................ 6-21
Figure 6.16 The TIC Chromatogram dialog ............................................................................... 6-24
Figure 6.17 The Analog Chromatogram dialog.......................................................................... 6-25
Figure 6.18 The Align Chromatogram Time dialog ................................................................... 6-26
Figure 6.19 Typical Chromatogram showing the Chromatogram Pointer.................................. 6-27
Figure 6.20 The Chromatogram Display Range dialog.............................................................. 6-28
Figure 6.21 The Centre Display dialog ...................................................................................... 6-28
Figure 6.22 The Center on Peak List dialog ............................................................................... 6-28
Figure 6.23 The Chromatogram Magnify dialog........................................................................ 6-29
Figure 6.24 The Default Chromatogram Range dialog .............................................................. 6-31
Figure 6.25 The Chromatogram Display View dialog ............................................................... 6-32
Figure 6.26 The Chromatogram Peak Annotation dialog........................................................... 6-35
Figure 6.27 The Short Cut Remove Chromatogram dialog........................................................ 6-36
Figure 6.28 The Remove Chromatogram dialog ........................................................................ 6-36
Figure 6.29 The Edit Text String dialog..................................................................................... 6-37
Figure 6.30 The Background Subtract dialog............................................................................. 6-39
Figure 6.31 The Background Subtract Status dialog .................................................................. 6-40
Figure 6.32 Unprocessed Total Ion Chromatogram ................................................................... 6-40
Figure 6.33 Background Subtracted Chromatogram produced using the parameters
shown in Figure 6.30............................................................................................... 6-40
Figure 6.34 Checking the operation of Background Subtract..................................................... 6-41
Figure 6.35 The Smooth chromatogram dialog.......................................................................... 6-42
Figure 6.36 Automatically setting the Window size (scans) ± parameter .................................. 6-42
Figure 6.37 Typical Smooth message box.................................................................................. 6-43
Figure 6.38 Results of chromatogram smoothing....................................................................... 6-43
Figure 6.39 The Integrate chromatogram dialog ........................................................................ 6-44
Figure 6.40 Setting the peak-to-peak noise amplitude ............................................................... 6-45
Figure 6.41 Peak Integration in Progress message ..................................................................... 6-46
Figure 6.42 The Peak Detect dialog ........................................................................................... 6-47
Figure 6.43 Join valleys parameter set to 15% ........................................................................... 6-48
Figure 6.44 Join valleys parameter set to 60% ........................................................................... 6-48
Figure 6.45 Reduce peak tailing parameter set to 150% ............................................................ 6-49
Figure 6.46 Reduce peak tailing parameter set to 50% .............................................................. 6-49
Figure 6.47 Raise baseline parameter set to 50% ....................................................................... 6-50
Figure 6.48 Raise baseline parameter set to 5% ......................................................................... 6-50
Figure 6.49 Draw verticals parameter set to 50%....................................................................... 6-51
Figure 6.50 Draw verticals parameter set to 95%....................................................................... 6-51
Figure 6.51 The ApexTrack Peak Detection Parameters dialog................................................. 6-52
Figure 6.52 Chromatogram Peak and Inverted Second Derivative ............................................ 6-53
Figure 6.53 The Response Threshold dialog .............................................................................. 6-53
Figure 6.54 The Edit Integrated Peaks dialog ............................................................................ 6-54
Figure 6.55 Peak Edit Error warning: Peak can’t overlap other peaks ....................................... 6-56
Figure 6.56 Peak Edit Error warning: Start and end positions are invalid.................................. 6-56
Figure 6.57 The Peak Purity dialog ............................................................................................ 6-57
Figure 6.58 Simple Peak Purity.................................................................................................. 6-58
Figure 6.59 Bayesian Peak Purity............................................................................................... 6-58
Figure 6.60 Signal to Noise processed chromatogram ............................................................... 6-60
Figure 6.61 The Signal to Noise dialog ...................................................................................... 6-60
Figure 6.62 The Auto Find Components dialog ......................................................................... 6-63
Figure 6.63 Mass Measure dialog parameters ............................................................................ 6-65
Figure 6.64 Component Worklist and annotated chromatogram................................................ 6-65
Figure 6.65 Combined spectra at scan 129 displaying component assignments.
Upper trace centroided data and lower trace continuum data.................................. 6-66
Figure 6.66 Component Worklist dialog displaying listing of component files ......................... 6-66
Figure 6.67 Mass Mapping component masses against the FASTA protein sequence database 6-68

Page 6-4
 Chromatogram

Figure 6.68 Mass Profile Fingerprint identifying beta-lactoglobulin as

top likelihood scoring protein ................................................................................. 6-69
Figure 6.69 The Edit Peak List dialog........................................................................................ 6-70
Figure 6.70 The File Open dialog .............................................................................................. 6-71
Figure 6.71 The Edit Peak List dialog for a new Peak List file.................................................. 6-71
Figure 6.72 The Get Peak List Entry dialog............................................................................... 6-73
Figure 6.73 MS/MS TIC chromatogram dropping to zero when the instrument
is in MS mode ......................................................................................................... 6-74
Figure 6.74 The Retention Index Table dialog........................................................................... 6-76
Figure 6.75 The Make Retention Index Calibration dialog........................................................ 6-77
Figure 6.76 Retention Index calibration curve ........................................................................... 6-78

Page 6-5
Chromatogram

Page 6-6
 Chromatogram

Getting Started
To Display the Total Ion Current (TIC) Chromatogram

Select the Sample List Menu Bar Chromatogram command; the Total Ion Current (TIC)
Chromatogram is displayed.

To Display a Summed Mass Chromatogram Around a Peak in a

Spectrum

Either:

Double-click on a peak in a Spectrum. The Summed Mass Chromatogram centered on

the selected peak and 1 Da wide is displayed.

Or:

1.
Select the Chromatogram Tool Bar button. The Mass Chromatogram dialog is
invoked.

2. Enter the required mass in the Description: text box.

3. Select the OK button. The Mass Chromatogram dialog is closed and the Summed
Mass Chromatogram is displayed.

The Chromatogram Window

Figure 6.1 The Chromatogram Window

Page 6-7
Chromatogram

The Chromatogram application runs in a top-level Window that has a Menu Bar and Tool Bar at
the top.

The Window may contain one or more Chromatogram Windows; each can contain one or more
Chromatogram traces.

When there is more than one trace in a Window, the current trace is identified by a colored square
at the left of the trace. To select another trace, click on any part of the required trace, or select a
trace from the Menu Bar Display, Traces command, or use the keyboard up and down arrow keys.

The chromatograms in each Chromatogram Window share a common time axis; place
Chromatograms in separate Windows to display them on different time axes.

The Chromatogram Menu Bar

The Chromatogram File Menu

Figure 6.2 The Chromatogram File Menu

Open Opens a data file.

Print Prints the current Window.

Printer Setup Invokes the standard Windows Print Setup dialog.

Exit Closes the Chromatogram window.

The Chromatogram Edit Menu

Figure 6.3 The Chromatogram Edit Menu

Copy Picture Copies the current Window to the clipboard.

Copy Copies a list of points in the chromatogram to the clipboard.

Chromatogram
List

Page 6-8
 Chromatogram

Copy Detected Copies a list of detected peaks to the clipboard.

Peaks
Paste Pastes the clipboard contents into the display.

Paste Special Invokes the standard Windows Paste Special dialog.

Peak List Read Invokes the Get Peak List Entry dialog, used to read a Peak List file into
the Chromatogram Window; see the “To Select a Peak List File” section,
on page 6-73.

Peak List Write Invokes the Edit Peak List dialog, used to add peak integration results to
any Peak List and edit Peak Lists; see the “To Create a New Peak List
File” section, on page 6-71.

Integrated Peaks Invokes the Edit Integrated Peaks dialog, used to edit integration results;
see the “Editing Integrated Peaks” section, on page 6-54.

The Chromatogram Display Menu

General

Figure 6.4 The Chromatogram Display Menu

Mass Invokes the Mass Chromatogram dialog, used to select a Mass

Chromatogram; see the “The Mass Chromatogram dialog” section, on
page 6-20.

Tic Invokes the TIC Chromatogram dialog, used to display a TIC or BPI
Chromatogram; see the “To Display a TIC or BPI Chromatogram using
the Menu” section, on page 6-24.

Page 6-9
Chromatogram

Analog Invokes the Analog Chromatogram dialog, used to select Analog Data
Channels for display; see the “To Display Analog Data Channels” section,
on page 6-25.

Remove Invokes the Remove Chromatogram dialog, used to remove multiple

Chromatogram traces from the display; see the “To Remove Multiple
Chromatogram Traces from the Display” section, on page 6-36.

Real-Time Toggles real time Chromatogram data update on and off.

Update
Range Invokes the Range sub-menu, see the “The Chromatogram Display, Range
Sub-menu” section, on page 6-11, for details.

Pointer Activates/deactivates the Chromatogram Pointer, see the “The

Chromatogram Pointer” section, on page 6-27, for details.

View Invokes the Chromatogram Display View dialog, used to edit the
Chromatogram Window Display Parameters; see the “To Change the
Display Parameters” section, on page 6-32.

Fraction Display Invokes the Fraction Display Parameters dialog, see the “FractionLynx
User’s Guide” for details.

Note:
This option only appears in the Chromatogram Display menu if the
FractionLynx Application Manager has been installed in MassLynx.

Peak Annotation Invokes the Chromatogram Peak Annotation dialog, used to edit the
Peak Annotation Parameters; see the “To Change the Peak Annotation
Parameters” section, on page 6-34.

Customize Invokes the Customize Toolbar dialog; see the “Customizing the
Toolbar Chromatogram Tool Bar” section, on page 6-16.

Toolbar Toggles the Tool Bar on and off.

Status bar Toggles the Status Bar on and off.

Move to Last Moves the currently selected Chromatogram to the top of the display, see
the “Changing the Order of Displayed Chromatograms” section, on
page 6-37, for further details.

Move to First Moves the currently selected Chromatogram to the bottom of the display,
see the “Changing the Order of Displayed Chromatograms” section, on
page 6-37, for further details.

Traces Displays a list of the Chromatograms in the display; click on a

Chromatogram in the list to select it.

Page 6-10
 Chromatogram

The Chromatogram Display, Range Sub-menu

Figure 6.5 The Chromatogram Display, Range Sub-menu

From Invokes the Chromatogram Display Range dialog, used to change the
horizontal axis range; see the “Altering the Range of the Horizontal Axis
using the Menu Bar” section, on page 6-28.

Default Invokes the Default Chromatogram Range dialog, to specify the default
horizontal axis range; see the “To Change the Default Display” section, on
page 6-31.

Magnify Invokes the Chromatogram Magnify dialog, used to magnify a section of

the current Chromatogram trace; see the “Setting Single or Multiple
Magnification Ranges using the Menu Bar Magnify Command” section,
on page 6-29.

Center Invokes the Center sub-menu.

On time or Invokes the Centre Display dialog, used to center the display around a
On scan point on the horizontal axis; see the “To Center the Display Around a
Point on the Horizontal Axis” section, on page 6-28.

Note:
This command changes depending on the units currently displayed on the
horizontal axis, the Centre Display dialog is invoked in either case.

Peak List Invokes the Center on Peak List dialog, used to center the display around
Entry a Peak List entry; see the “To Center the Display Around a Peak List
Entry” section, on page 6-28.

Page 6-11
Chromatogram

Figure 6.6 The Chromatogram Display, Range, Center sub-menu

Align Invokes the Align Chromatogram Time dialog, used to align two
Chromatogram traces in time; see the “To Align Two Chromatograms”
section, on page 6-26.

The Chromatogram Process Menu

Figure 6.7 The Chromatogram Process Menu

Integrate Invokes the Integrate chromatogram dialog, see the “To Integrate a
Chromatogram” section, on page 6-44.

Purity Invokes the Peak Purity dialog; which processes TIC Chromatograms that
have been integrated, see the “Peak Purity” section, on page 6-57.

Smooth Invokes the Smooth chromatogram dialog; see the “The Smooth
Chromatogram Dialog” section, on page 6-41.

Subtract Invokes the Background Subtract dialog; see the “Performing a

Background Subtract” section, on page 6-39.

Process All Press once to process all traces in the current window. Press again to
Traces process only the current trace in the current window.

Page 6-12
 Chromatogram

Combine Spectra Invokes the Combine Spectrum dialog, used to combine spectrum scans
across a chromatogram peak; see the “Combine Spectra” section in
Chapter 7, “Spectrum” for further details.

Components Invokes the Components sub-menu.

Auto Find Invokes the Auto Find Components dialog, used to find all multiply-
charged components from all scans in the current peak detected
chromatogram; see the “Automatic Component Finding” section, on
page 6-64.

Edit Worklist Invokes the Component Worklist dialog, used to analyze and edit the
component list; see the “The Component Worklist Dialog” section, on
page 6-66.

Figure 6.8 The Process, Components sub-menu

Signal to Noise Invokes the Signal To Noise dialog, used to calculate the Signal To Noise
value for a mass chromatogram; see the “Signal to Noise Ratio” section,
on page 6-59.

The Chromatogram Window Menu

Figure 6.9 The Chromatogram Window Menu

Tile Displays the current windows in a tiled view.

Cascade Displays the current windows in a cascaded view.

Stack Displays the current windows in a stacked view.

Arrange Icons Arranges the icons of minimized windows at the bottom of the
Chromatogram Window.

Page 6-13
Chromatogram

New Trace Invokes the New Chromatogram dialog see the “The New
Chromatogram Dialog” section, on page 6-20, for details.

List of current Click on the required trace to select it.

traces

The Chromatogram Tools Menu

Figure 6.10 The Chromatogram Tools Menu

Retention Index Invokes a sub-menu with the following items.

Edit Index Invokes the Retention Index Table dialog; this allows the Retention
Table Index Table to be edited, see the “The Retention Index Table dialog”
section, on page 6-76.

Make Invokes the Make Retention Index Calibration dialog; see the “To Make
Calibration a Retention Index Calibration” section, on page 6-77.

Delete Deletes the Retention Index Calibration, if present. A warning window is

Calibration invoked.

Calibration Invokes the Retention Index Status dialog; this displays details of the
Status Retention Index Calibration status.

Figure 6.11 Typical Retention Index Status dialog

The Chromatogram Help Menu

The Help, Chromatogram command invokes the Help function for Chromatogram.

Page 6-14
 Chromatogram

The Chromatogram Tool Bar

General

The Chromatogram Tool Bar is displayed at the top of the Chromatogram Window and allows the
User to perform common operations with a single click of the appropriate Tool Bar button. The
default Chromatogram Tool Bar contains the buttons listed below. The Tool Bar may be
customized and additional buttons displayed for other Chromatogram operations, see the
“Customizing the Chromatogram Tool Bar” section, on page 6-16.

Tool Menu equivalent Purpose

Bar
button
File, Open Opens a data file.

File, Print Prints the current Window in portrait format.

File, Print Prints the current Window in landscape format.

Edit, Copy Picture Copies the current Window to the clipboard.

Edit, Copy Copies a list of points in the chromatogram to the

Chromatogram List clipboard.

Edit, Copy Detected Copies a list of detected peaks to the clipboard.

Peaks
Edit, Paste Pastes the clipboard contents into the display.

Display, Mass Invokes the Mass Chromatogram dialog; this is used

to select a Mass Chromatogram, see the “The Mass
Chromatogram dialog” section, on page 6-20.

Process, Integrate Performs peak integration, see the “To Integrate a

Chromatogram” section, on page 6-44.

Process, Combine Invokes the Combine Spectrum dialog; used to

Spectra combine spectrum scans across a chromatogram peak;
see the “Combine Spectra” section in Chapter 7,
“Spectrum” for further details.

Process, Process All Toggles between processing all traces in the current and
Traces processing only the current trace in the current window.

Invokes the Edit Text String dialog; this allows text to

be added to a Chromatogram.

Toggles between each subsequent chromatogram, or

chromatogram process, appearing in a new window and
being added to the current one.

Pressing once causes each subsequent chromatogram or

chromatogram process to replace the currently selected
trace. Pressing a second time cancels this mode.

Page 6-15
Chromatogram

Note:

1. The button is grayed when the button is depressed.

2. The manner in which chromatograms are added to the Chromatogram Window can also be
selected via the Menu Bar Window, New Trace command, refer to the “The New
Chromatogram Dialog” section, on page 6-20, for details.

Display, Real-Time Toggles real time Chromatogram data update on and

Update off.

Display, Range, Increases the magnification of the current range.

Magnify
Display, Range, Decreases the magnification of the current range.
Magnify
Display, Range, Deletes the current magnification range.
Magnify
Resets the display to a TIC trace.

Decrements the currently displayed scan in the

associated Spectrum window.

Increments the currently displayed scan in the

associated Spectrum window.

Press once to restore the previous display range; press

again to use the default display range.

Customizing the Chromatogram Tool Bar

General

The Chromatogram Tool Bar can be customized to:

• Add buttons for frequently used operations.

• Remove buttons that are not required.

• Change the order in which the Tool Bar buttons are displayed.

The additional buttons that can be added to the default Chromatogram Tool Bar are:

Tool Menu equivalent Purpose

Bar
button
Edit, Integrated Peaks Invokes the Edit Integrated Peaks dialog.

Display, Analog Invokes the Analog Chromatogram dialog.

Process, Smooth Invokes the Smooth chromatogram dialog.

Process, Subtract Invokes the Background Subtract dialog.

Page 6-16
 Chromatogram

Window, Tile Displays the current windows in a tiled view.

Window, Cascade Displays the current windows in a cascaded view.

Window, Stack Displays the current windows in a stack view.

The Customize Toolbar dialog

To customize the Chromatogram Tool Bar, select the Chromatogram Menu Bar Display,
Customize Toolbar command; the Customize Toolbar dialog is invoked.

Figure 6.12 The Customize Toolbar dialog

Available This list box contains all the available buttons that are not currently in the
Buttons: list box Tool Bar. A button can be selected by clicking on it.

The top entry in the box is Separator; it is never removed from the
Available Buttons: list box, although it can be added to the Toolbar
Buttons list box and thus insert a separation gap between the buttons in
the Tool Bar.

Tool Bar This list box contains all the buttons that are currently in the toolbar. A
Buttons: list box button can be selected by clicking on it. The last entry in this box is
always Separator (dimmed); it cannot be removed from the list box, it
allows buttons to be added to the end of the list.

Add-> Moves the selected button from the Available Buttons: list box to the
Tool Bar Buttons: list box.

<-Remove Moves the selected button from the Tool Bar Buttons: list box to the
Available Buttons: list box.

Note:
This push-button is grayed if no item is selected in the Tool Bar Buttons:
list box.

Close Exits the Customize Toolbar dialog.

Reset Resets the Tool Bar to its default display.

Page 6-17
Chromatogram

Move Up Moves the selected button one position up the list in the Toolbar Buttons:
list box.

Note:
This push-button is grayed if no item is selected in the Tool Bar Buttons:
list box, or if the top item in the list is selected.

Move Down Moves the selected button one position down the list in the Toolbar
Buttons: list box.

Note:
This push-button is grayed if no item is selected in the Tool Bar Buttons:
list box, or if the bottom item in the list is selected.

To Add Buttons to the Tool Bar

1. Select the Chromatogram Menu Bar Display, Customize Toolbar command; the Customize
Toolbar dialog is invoked.

2. Select the button to be added in the Available Buttons: list box.

3. Select the Tool Bar button before which the new button is to be added in the Toolbar
Buttons: list box.

4. Select the Add button. The new button is added to the Toolbar Buttons: list box.

5. Repeat steps 2 to 4 to add further buttons to the Tool Bar.

6. Separators can be inserted between Tool Bar buttons to divide them into logical groups. To
add a separator, repeat steps 2 to 4 selecting Separator in the Available Buttons: list box.

7. Select the Close button to exit and save the changes.

To Remove Buttons from the Tool Bar

1. Select the Chromatogram Menu Bar Display, Customize Toolbar command; the Customize
Toolbar dialog is invoked.

2. Select the button to be removed in the Toolbar Buttons: list box.

3. Select the Remove button. The button is removed from the Toolbar Buttons: list box.

4. Repeat steps 2 and 3 to remove further buttons from the Tool Bar.

5. Select the Close button to exit and save the changes.

To Change the Order in which Tool Bar Buttons are Displayed

1. Select the Chromatogram Menu Bar Display, Customize Toolbar command; the Customize
Toolbar dialog is invoked.

2. Select the button to be moved in the Toolbar Buttons: list box.

3. Select the Move Up or Move Down buttons to move the Tool Bar button.

4. Repeat steps 2 and 3 as often as required.

5. Select the Close button to exit and save the changes.

Page 6-18
 Chromatogram

To Reset the Tool Bar to the Default Settings

1. Select the Chromatogram Menu Bar Display, Customize Toolbar command; the Customize
Toolbar dialog is invoked.

2. Select the Reset button.

3. Select the Close button to exit and save the changes.

To Remove the Tool Bar from the Chromatogram Display

Select the Menu Bar Display, Toolbar command, the Tool Bar is removed from the display. A
tick mark appears next to this menu item when it has been selected.

To re-display the Tool Bar, select the Menu Bar Display, Toolbar command again.

Displaying Chromatograms
Adding or Replacing Chromatogram Traces

MassLynx provides a number of options for displaying new chromatogram traces. New
chromatogram traces can be generated by:

• Opening a new file.

• Processing Chromatogram traces (subtract, smooth, integrate, etc.).

• Selecting mass Chromatograms by double-clicking on a spectrum, or by using the Menu Bar

Display, Mass command.

To display each new chromatogram trace in a new window, select the Tool Bar button. To
cancel this mode and display new traces in the existing window select the Tool Bar button
again.

When a new trace is displayed in the existing window, it can be added to the traces currently
displayed, or it can replace the current trace. Select the Tool Bar button once to cause each
subsequent chromatogram, or chromatogram process, to replace the currently selected trace.
Selecting the button a second time causes each subsequent chromatogram or chromatogram
process to be added to the traces on display. Up to sixteen chromatogram traces can be displayed
in one window.

Note:

1. The button is grayed when the button is depressed.

2. The manner in which chromatograms are added to the Chromatogram Window can also be
selected via the Menu Bar Window, New Trace command, refer to the “The New
Chromatogram Dialog” section, on page 6-20, for details.

Page 6-19
Chromatogram

The New Chromatogram Dialog

The New Chromatogram dialog is used to select the manner in which chromatograms are added
to the Chromatogram Window; it is invoked by the Menu Bar Window, New Trace command.

Figure 6.13 The New Chromatogram dialog

Add Trace Adds the chromatogram to the current Chromatogram Window.

Replace Trace The chromatogram replaces the currently selected chromatogram in the
Chromatogram Window.

Replace All The chromatogram replaces all the chromatograms in the Chromatogram
Window.

New Window Displays the chromatogram in a new Window.

The Mass Chromatogram dialog

General

The Mass Chromatogram dialog is used to control the way in which summed mass
chromatograms are displayed. To invoke the Mass Chromatogram dialog, either:

Select the Menu Bar Display, Mass command.

Or:

Select the Tool Bar button.

In either case, the Mass Chromatogram dialog is invoked.

Note:
The Mass Chromatogram dialog format depends on the selected data file.

Page 6-20
 Chromatogram

Figure 6.14 The Mass Chromatogram dialog for full scan data

Figure 6.15 The Mass Chromatogram dialog for Selected Ion Recording (SIR) data

File: field Displays the name of the current data file.

Description: A description of the mass chromatogram to be generated can be entered in

this text box

Note:
1. x, y and z represent either masses or channels, depending on the data
file type.

2. For a SIR data file, the required channels can be entered by double-
clicking on the appropriate channels in the Channels: list box, see
below. They can also be entered by typing directly into the
Description: text box, using the format n, or Chn, where n is the
channel identifier, as displayed in the Channels: list box.

3. For a full-scan data file, the required masses can be entered by typing
the numbers into the box. They can also be entered by right-clicking
in any Spectrum Window. The peak closest to the cursor's mass
position having an intensity greater than, or equal to, the cursor
height is selected. A right click-and-drag operation can be used to
enter a range of masses. Commas automatically separate multiple
mouse selections.

Page 6-21
Chromatogram

The formats for entries in this box are as follows:

x Generates the chromatogram of x.

x+y Generates the chromatogram of x added to y.

x-y Generates the chromatogram of y subtracted from x.

x_y Generates the chromatogram of the summation of all

masses/channels from x to y.

Note:
More than one mass chromatogram trace can be generated simultaneously
by separating individual descriptions with commas. e.g.:

x,y Generates the two chromatograms centered around x and y.

x_y,z Generates the chromatogram of the summation of all

masses/channels from x to y and the chromatogram of z.

Examples of these formats are:

110 The summed chromatogram of masses 109.5 to 110.5.

110+340 The summed chromatogram of masses 109.5 to 110.5 and

339.5 to 340.5.

110-340 The summed chromatogram of masses 339.5 to 340.5

subtracted from the summed chromatogram of masses 109.5 to
110.5.

110_340 The summed chromatogram of all masses from 110 to 340

inclusive.

110, 150 The two mass chromatograms centered around 110 and 150.

The summed mass chromatogram of all masses from 110 to

110_150, 150, and the mass chromatogram centered around 340.
340
Function: Presents a list of the functions associated with the data file. The User can
select the required function with the mouse. If the function is changed the
Description: box is blanked.

Channels: (SIR data files only). The channel masses of the selected Function are
listed with ascending order, numeric identifiers.

Add trace Adds the chromatogram to the current Chromatogram Window.

Replace trace The chromatogram replaces the currently selected chromatogram in the
Chromatogram Window.

New window Displays the chromatogram in a new Window.

Page 6-22
 Chromatogram

OK Confirms the selections in the Description: box and produces the required
chromatograms. The Mass Chromatogram dialog is closed. The chosen
button (Add trace, Replace trace, or New window) is retained as the
default button. The OK button is grayed if the Description: box is empty.

File Invokes the Chromatogram Data Browser dialog, see Chapter 3, “The
MassLynx Window and Related Information” for further information.

Select All (SIR data files only). Enters all the channels in the Channels: box into the
Description: box, separated by commas.

To Display a Summed Mass Chromatogram

1. Select the Chromatogram Tool Bar button, or select the Chromatogram Menu Bar
Display, Mass command, the Mass Chromatogram dialog is invoked.

2. If required, select a function from the Function: list box.

3. Enter the description of the mass chromatogram to be generated in the Description: text box;
refer to the “General” section, on page 6-20, for formatting details.

4. If the Mass Chromatogram is to be added to the current Chromatogram Window, select the
Add trace option. If the Mass Chromatogram is to replace the current Chromatogram
Window, select the Replace trace option. If the Mass Chromatogram is to be placed in a new
Chromatogram Window, select the New window option.

5. Select the OK button.

Generating Mass Chromatograms from a Spectrum Display

Mass Chromatograms can be generated by right-clicking in any Spectrum display. The peak
closest to the cursor's mass position having an intensity greater than, or equal to, the cursor height
is selected.

A right-click and drag operation generates a chromatogram for the selected range.

Note:
The new Chromatogram will be added to the current window, replace a trace in the current
window, or be placed in a new window depending on the current setting in the Mass
Chromatogram dialog.

To Display an Accurate Mass Chromatogram

1. Select the MassLynx Tools Shortcut Bar, Options icon; the Options dialog is invoked, see
Chapter 3, “The MassLynx Window and Related Information”.

2. In the Mass Chromatogram Window frame, select the required option [Parts per million or
Abs window (Da)] and enter an appropriate value in the adjacent text box.

3. Select the OK button; the Options dialog is closed.

4. Select the Chromatogram Tool Bar button, or select the Chromatogram Menu Bar
Display, Mass command; the Mass Chromatogram dialog is invoked.

5. Enter the mass to the required accuracy (up to four decimal places) in the Description: box.

6. Select the OK button.

Page 6-23
Chromatogram

To Display the Mass Chromatograms for a New Data File

1. Select the Chromatogram Menu Bar File, Open command; the Chromatogram Data
Browser is invoked.

2. Select the new data file to be displayed.

3. Select the Replace All option. This will replace the existing data file and any Mass
Chromatograms that are on display.

4. Select the OK button.

TIC and BPI Chromatograms

General

The Total Ion Current (TIC) Chromatogram is the default chromatogram displayed when the
Chromatogram application is started, or when a new data file is selected using the File, Open
command. The intensity plotted at each point in the TIC is the sum of all the intensities in that
scan. The TIC Chromatogram can also be obtained by selecting the Tool Bar button.

A Base Peak Intensity (BPI) Chromatogram plots the greatest intensity at each scan whereas the
TIC is the sum of the noise and signal at each scan. The BPI Chromatogram exhibits a greater
apparent resolution and signal-to-noise but will only contain contributions from the most intense
components. Therefore, it is possible that some peaks in the TIC Chromatogram may not be
visible in the BPI Chromatogram.

To Display a TIC Chromatogram using the Tool Bar

Select the Tool Bar button. The Chromatogram display will be updated to show a single TIC
Chromatogram for the currently selected trace.

To Display a TIC or BPI Chromatogram using the Menu Bar

1. Select the Chromatogram Menu Bar Display, TIC command. The TIC Chromatogram
dialog is invoked.

Figure 6.16 The TIC Chromatogram dialog

Page 6-24
 Chromatogram

2. Select the required function in the Function: list box.

3. If a BPI Chromatogram is required, select the BPI Chromatogram box.

4. If the Chromatogram is to be added to the current Chromatogram Window, select the Add
trace option. If the Chromatogram is to replace the current trace, select the Replace trace
option. If the Chromatogram is to have its own Window, select the New window option.

5. Select the OK button.

Analog Chromatograms

General

During an acquisition, MassLynx can store analog information obtained from an auxiliary source
such as a UV detector. The acquisition may be set up to include analog data by selecting the
Analog Channel controls in the Analog Data dialog; see the appropriate Instrument User’s Guide
for details. Up to four channels of analog data may be acquired.

To Display Analog Data Channels

1. Select the Chromatogram Menu Bar Display, Analog command; the Analog Chromatogram
dialog is invoked.

Figure 6.17 The Analog Chromatogram dialog

2. Select the required trace from the Channel: list box. If the list box is empty, the acquisition
was not set to include analog data.

3. If the Chromatogram is to be added to the current Chromatogram Window, select the Add
trace option. If the Chromatogram is to replace the current trace, select the Replace trace
option. If the Chromatogram is to have its own Window, select the New window option.

4. If the trace is to be aligned with an existing trace, select the Align button; the Align
chromatogram Time dialog is invoked, see the “To Align Two Chromatograms” section, on
page 6-26, for details.

5. Select the OK button.

Page 6-25
Chromatogram

Aligning Analog Chromatograms

General

Data from the auxiliary detector may be slightly out of phase with data from the chromatography
system as there may be a time lag between the sample arriving at the auxiliary detector and at the
chromatography system.

An offset to the time axis of each analog trace can be specified to allow it to be manually aligned
with another. A different time offset may be applied to each of the analog channels acquired.
Only the display is affected; the data on disk remains unchanged.

Note:
This only works if the horizontal axis is displayed as “time” and not “scans”.

To Align Two Chromatograms

1. Select the Chromatogram Menu Bar Display, Analog command. The Analog
Chromatogram dialog is invoked.

2. Select the Align button; the Align chromatogram Time dialog is invoked.

Note:
The Align Chromatogram Time dialog is also invoked by the Menu Bar Display, Range, Align
command.

Figure 6.18 The Align Chromatogram Time dialog

3. Enter the Offset time that is required to line up the two chromatograms

4. Select the OK. button; the Align Chromatogram Time dialog is closed.

Page 6-26
 Chromatogram

The Chromatogram Pointer

The Chromatogram Pointer is used to identify spectra for the Spectrum service to display. It is
activated/deactivated by the Menu Bar Display, Pointer command.

Figure 6.19 Typical Chromatogram showing the Chromatogram Pointer

Double-clicking with the mouse on the chromatogram trace moves the Pointer to the mouse
position and activates a linked Spectrum window. Right-clicking with the mouse also has this
effect, but doesn’t activate the Spectrum window.

The pointer can also be dragged, using its triangular top, to the required position on the
chromatogram trace.

The linked Spectrum window is updated to reflect the new pointer position whatever the method
used.

Manipulating the Display

Altering the Range of the Horizontal Axis

Altering the Range of the Horizontal Axis using the Mouse

Click and hold the left mouse button at one end of the region of interest and drag the cursor
horizontally to the other end. As the cursor is dragged a “rubber band” is stretched out to indicate
the range selected; do not go beyond the bounds of the axis. When the mouse button is released
the selected range will be re-displayed to fill the current window.

Page 6-27
Chromatogram

Altering the Range of the Horizontal Axis using the Menu Bar

1. Select the Menu Bar Display, Range, From command; the Chromatogram Display Range
dialog is invoked.

Figure 6.20 The Chromatogram Display Range dialog

2. Enter new From and To values for the horizontal axis.

3. Select the OK button.

Centering the Display Around a Particular Point

To Center the Display Around a Point on the Horizontal Axis

1. Select the Menu Bar Display, Range, Center, On time or Display, Range, Center, On Scan
command, as appropriate (only one of these options will be on the menu, depending on the
units currently displayed on the horizontal axis). The Centre Display dialog is invoked.

Figure 6.21 The Centre Display dialog

2. Specify the scan number or retention time to Center on.

3. Specify the half-width of the display range in the Window text box.

4. Select the OK button.

To Center the Display Around a Peak List Entry

1. Select the Menu Bar Chromatogram Display, Range, Center, Peak List Entry command; the
Center on Peak List dialog is invoked.

Figure 6.22 The Center on Peak List dialog

2. Specify the Peak List Entry to center on.

3. Specify the half-width of the display range in the Window text box.

Page 6-28
 Chromatogram

4. Select the OK button.

Altering the Range of the Intensity Axis

Click and hold the left mouse button at one end of the region of interest and drag the cursor
vertically to the other end. As the cursor is dragged a “rubber band” is stretched out to indicate the
range selected; do not go beyond the bounds of the axis. When the mouse button is released the
selected range will be re-displayed to fill the current window.

This operation can be repeated as often as required.

Altering the Range of Both Axes

Click and hold the left mouse button at one end of the region of interest and drag the cursor to the
diagonally opposite corner. As the cursor is dragged a “rubber band” is stretched out to indicate
the region selected; do not go beyond the bounds of the axes. When the mouse button is released
the selected region will be re-displayed to fill the current window.

This operation can be repeated as often as required.

Setting Magnified Ranges

Setting Single or Multiple Magnification Ranges using the Mouse

Click and hold the middle mouse button at one end of the region of interest and drag the cursor
horizontally to the other end. As the cursor is dragged a “rubber band” is stretched out to indicate
the range selected; do not go beyond the bounds of the axis. When the mouse button is released
the selected range will be re-displayed with an initial magnification factor of two.

Alternatively, pressing the Shift key while using the left mouse button will perform the same
operation.

Setting Single or Multiple Magnification Ranges using the Menu Bar Magnify Command

1. Either:

a. Select the Menu Bar Display, Range, Magnify command.

Or:

b. Double-click on the range magnification description of an existing magnified range.

In either case, the Chromatogram Magnify dialog is invoked.

Figure 6.23 The Chromatogram Magnify dialog

Page 6-29
Chromatogram

2. Enter the magnification factor to be applied in the By text box.

3. Enter the range to be magnified in the From and To text boxes.

4. To define more than one magnification range on the displayed Chromatogram, select a new
range in the Range list box and repeat Steps 2 and 3. Up to five different magnified regions
of the Chromatogram can be defined.

5. Select the OK button to close the dialog. The Chromatogram is re-displayed with the data in
the selected regions magnified by the requested factor. The magnified regions are displayed
in a different color and labeled with the magnification factor.

Where multiple magnification regions have been defined, to select the current magnification range,
click in the magnification description that appears above the range. The description will change
color to red, to indicate the currently selected range.

Setting the Intensity Axis Magnification Range using the Tool Bar

Select to increase the magnification of the current range. The current magnification
factor is multiplied by 1.5, and rounded up to the nearest even number to give the
increased magnification factor. For example, if the initial magnification factor is 2, this
will give subsequent magnification factors of 4, 6, 10, 16, etc.

Select to decrease the magnification of the current range. The current magnification
factor is divided by 1.5, and rounded down to the nearest even number to give the
decreased magnification factor. For example, if the initial magnification factor is 16, this
will give subsequent magnification factors of 10, 6, 4, etc.

To Change the Magnification of a Particular Range

1. Either:

a. Select the Menu Bar Display, Range, Magnify command.

Or:

b. Double-click on the range magnification description of an existing magnified range.

In either case, the Chromatogram Magnify dialog is invoked, see the “Setting Single or
Multiple Magnification Ranges using the Menu Bar Magnify Command” section, on
page 6-29.

2. Enter the new magnification factor in the By text box.

3. Select the OK button.

Deleting Magnification Ranges

Select the Tool Bar button to delete the current magnification range.

To delete all the magnification ranges:

1. Either:

a. Select the Menu Bar Display, Range, Magnify command.

Or:

Page 6-30
 Chromatogram

b. Double-click on the range magnification description of an existing magnified range.

In either case, the Chromatogram Magnify dialog is invoked, see the “Setting Single or
Multiple Magnification Ranges using the Menu Bar Magnify Command” section, on
page 6-29.

2. Select the Default button; this will delete all magnification ranges.

3. Select the OK button.

Restoring the Display

Selecting the Tool Bar button once restores the display to its previous state. Selecting it a
second time restores the display to the default range.

Note:
These operations do not remove magnification ranges.

Setting the Display Range Defaults

Note:

The display range default settings specify both the effects of selecting the Tool Bar button, and
adding a new Chromatogram to the display.

To Change the Default Display

1. Select the Menu Bar Display, Range, Default command; the Default Chromatogram Range
dialog is invoked.

2. Make the required changes, see below.

3. Select the OK button.

Figure 6.24 The Default Chromatogram Range dialog

Default graph If there is more than one chromatogram in a window, this option specifies
Frame whether the default time/scan range for that window is made large enough
to include the time/scan ranges of All the chromatograms, or large enough
for the Current chromatogram only.

Automatic range If this option is selected, the display range will return to the specified
default default (see Default graph above) when a new chromatogram is added to
a Chromatogram Window. If this option is not checked, the display range
will remain unchanged when a new chromatogram is added.

Page 6-31
Chromatogram

Controlling the Appearance of the Display

General

Each Chromatogram Window has its own set of Display Parameters, which determine the
appearance of the Chromatogram display. The Parameters can be inspected and altered for the
current Chromatogram Window from the Chromatogram Display View dialog.

To Change the Display Parameters

1. Select the Menu Bar Display, View command; the Chromatogram Display View dialog is
invoked.

2. Make the required changes, see below.

3. Select the OK button.

Figure 6.25 The Chromatogram Display View dialog

Normalize Data These controls specify the scale on the intensity axis.
To Frame
Largest Peak Displays the largest peak at 100% of the intensity axis.

Intensity When selected, 100% on the intensity axis represents the intensity
specified in the adjacent text box.

Baseline at Scales the intensity axis so that the baseline is at zero intensity.
Zero
Baseline abs Scales the intensity axis so that the baseline is at the intensity specified in
the adjacent text box.

Page 6-32
 Chromatogram

Baseline % Moves the baseline to the percentage value entered in the text box; this
value is a percentage of the normalization intensity. The normalization
intensity is the largest peak intensity, unless the Intensity option is
selected, when it becomes the intensity specified by the Intensity edit box.

Lowest Point Automatically scales the display so that the lowest point of the trace is at
the bottom. This can be useful for displaying Diode Array data if the trace
has dropped below zero and the data has negative values.

Link Vertical Gives all axes in the current window a common vertical scale. This
Axes enables two chromatograms to be plotted on the same intensity scale, in
order to overlay and compare them.

Axis Label Frame

Horizontal Select the units for the horizontal axis from the list box; the options are
Axis retention Time and Scan number.

Style Frame

Overlay Allows multiple traces in the same window to be superimposed on the

Graphs same axis.

If the option is not selected, the traces will be drawn on separate axes,
arranged vertically.

Note:
When Chromatograms are overlaid, only the currently selected trace is
annotated.

Fill Trace Colors the area under the chromatogram.

Fill Detected Colors peaks detected by integration.

Peaks
Peak List For peak detected data only, the Time, Height, Area and Percentage Area
for each peak are listed on the right-hand-side of the Chromatogram.

Graph Header Displays header information at the top of the Chromatogram.

Process Displays process information in the Chromatogram header.

Description
Note:
The Graph Header option overrides the Process Description option, i.e.
if the Graph Header is deselected, the Process Description will also be
deselected.

Component For non-GC installations, displays a summary of the components

Table identified so far on each chromatogram.

Page 6-33
Chromatogram

Split Axis This option is enabled when the Overlay Graphs control is selected. It
allows the User to alter the aspect ratio of the chromatogram by dividing
the horizontal axis into segments, then arranging the segments vertically.
For example, if a chromatogram of 30 minutes duration is on display, and
3 is selected in the Split Axis option, the display will show three axes, one
from 0 to 10 minutes, one from 10 to 20 minutes, and one from 20 to
30 minutes.

Overlay This option is enabled when the Overlay Graphs control is selected. It
Step (%) allows the User to offset each subsequent chromatogram trace by a
percentage of the intensity axis. This can make it easier to examine
overlaid traces.

Grid Enables the User to specify a grid to be displayed on the Chromatogram

display. The pattern of the lines that make up the grid can be chosen as
Dot, Dash or Solid. Select Off if no grid is to be displayed.

Header button Invokes the Header Editor, which allows editing of the header
information displayed at the top of the window. For more information see
the “The Header Editor” section in Chapter 3, “The MassLynx Window
and Related Information”.

Controlling the Appearance of Peak Labels

General

Each Chromatogram Window has its own set of Peak Annotation Parameters, which determine
the appearance of peak labels. The User can inspect and alter the parameters for the current
window in the Chromatogram Peak Annotation dialog.

To Change the Peak Annotation Parameters

1. Select the Menu Bar Display, Peak Annotation command; the Chromatogram Peak
Annotation dialog is invoked.

2. Make the required changes, see below.

3. Select the OK button.

Annotation Type The parameters in this frame control the types of peak annotation that will
Frame appear on the chromatogram.

Peak Top Time Annotates peaks with their retention time values.

Peak Top Scan Annotates peaks with their corresponding scan numbers.

Peak Purity If the Peak Purity Process has been run, annotates peaks with the
calculated purity value to the number of decimal places specified in the
adjacent Decimal Places text box.

Page 6-34
 Chromatogram

Figure 6.26 The Chromatogram Peak Annotation dialog

Scan Base Annotates peaks with the base peak mass at that particular scan number to
Peak Mass the number of decimal places specified in the adjacent Decimal Places
text box.

Peak Response If peak detection has been performed, annotates peaks with their calculated
Area response areas to the number of decimal places specified in the adjacent
Decimal Places text box.

Peak Response Annotates peaks with their calculated peak heights.

Height

Annotation The parameters in this frame control the intensity thresholds for peak
Threshold Frame annotation.

% Full Scale When selected, only those peaks greater than the percentage of the current
base peak intensity specified in the adjacent text box will be annotated.

Intensity When selected, only those peaks greater than the absolute intensity value
specified in the adjacent text box will be annotated.

All Peaks Annotates all peaks, regardless of intensity.

Level Select High, Medium or Low from the list box, to determine the number
of labels to be displayed on the chromatogram.

BioLynx Frame This frame contains controls that are applicable to ElectroSpray data.

Component (Non-GC installations only.) Labels peaks with the name of the
Label appropriate component.

Digest Label (Non-GC installations only.) Labels the chromatogram with any digest
labels generated in BioLynx.

Page 6-35
Chromatogram

Scan Set Mass (Q-TOF data only). Annotates the peak with the set mass of the scan it
represents, to the number of decimal places specified in the adjacent
Decimal Places text box.

Removing Chromatograms from the Display

To Remove a Single Chromatogram Trace from the Display

1. Press the keyboard Delete key. A dialog is invoked asking for confirmation of deletion of the
currently selected chromatogram trace.

Figure 6.27 The Short Cut Remove Chromatogram dialog

2. Select the OK button; the dialog is closed and the selected traces are removed from the
display. This operation does not affect the data stored on disk.

To Remove Multiple Chromatogram Traces from the Display

1. Select the Chromatogram Menu Bar Display, Remove command; the Remove
Chromatogram dialog is invoked.

Figure 6.28 The Remove Chromatogram dialog

2. The traces in the current Window are listed in the order in which they appear on the display.
Select one or more traces in the list box by clicking on them. Clicking again on a selected
trace will cancel the selection. Select the All button to select all the traces.

3. Select the OK button; the dialog is closed and the selected trace(s) are removed from the
display. This operation does not affect the data stored on disk.

Real-Time Display of Chromatograms

If data are being acquired into a file, the associated chromatograms can be displayed in real time,
by selecting the Tool Bar button or the Menu Bar Display, Real-Time Update command.

Each Chromatogram Window has a separate real time update switch. The state of the switch for a
particular Window can be ascertained by checking if the Tool Bar button is depressed, or by
making that Window current, then selecting the Menu Bar Display menu. If real time update is
enabled, the Real-Time Update item has a tick mark by it.

Page 6-36
 Chromatogram

Changing the Order of Displayed Chromatograms

When a Window contains multiple traces, the order in which they are displayed can be changed.
The Chromatogram which is first in the list is displayed at the bottom of the screen, or, if graphs
are overlaid, on top of the others.

Select the Menu Bar Display, Move To First option to display the currently selected
Chromatogram at the bottom of the Window.

Select the Menu Bar Display, Move To Last option to display the currently selected
Chromatogram at the top of the Window.

Adding Text to the Chromatogram Display

To add user text labels to the chromatogram display:

1. Select the Tool Bar button.

2. Move the mouse cursor to the position where user text is required and click the button; the
Edit Text String dialog is invoked.

3. Enter the text in the Text window.

4. Select the desired options (see below).

5. Select the OK button.

The text’s position can be changed by clicking and dragging it to a new position. The text size can
be changed by clicking on it and dragging one of the handle boxes. To edit the text, double-click
on it to re-invoke the Edit Text String dialog.

The font and color of the user text can be changed using the MassLynx Tools Shortcut Bar Colors
and Fonts icon, which invokes the Colors and Fonts dialog, see Chapter 3, “The MassLynx
Window and Related information”. Any changes made will only apply to text added after the
changes. If the fonts or colors of existing text are to be changed, it must be deleted and reinserted.

Figure 6.29 The Edit Text String dialog

Page 6-37
Chromatogram

Border Displays a box around the user text.

Vertical Displays text vertically, rather than horizontally.

Autosize Select this option to automatically size the text area that holds the user
text. If it is not checked two handle boxes will appear on the screen, click
on one of them and drag until the text area is the required size.

Attach to axis Select this option to specify that text can only be positioned within a box
defined by the intensity and time/scan axes. If it is not selected, text can
be positioned anywhere on the screen.

Justification Text can be aligned to the Left, Center or Right of the text area.
Frame
The current formatting options are saved as the default options each time the Edit Text String
dialog is closed.

To delete user text from the display, click on it, then press the keyboard Delete key.

Processing Chromatograms
General

Three processes are available for use on chromatograms:

• Polynomial Background Subtraction (Background Subtract), see the “Background Subtract”

section, on page 6-39.

• Smoothing, see the “Smoothing Chromatograms” section, on page 6-41.

• Integration, see the “Integrating Chromatograms” section, on page 6-43.

Background Subtraction and Smoothing help to improve the presentation of the data. Integration
locates peaks, positions baselines and calculates peak statistics for quantitative work.

Processing Multiple Chromatograms

The Background Subtract, Smoothing and Integration processes can be performed automatically
on all the Chromatograms within the current Window. To enable this, select the Tool Bar
button, or select the Menu Bar Process, Process All Traces command; this menu item will have a
tick next to it when selected. To turn off multiple processing, reselect the Tool Bar button, or
the Menu Bar Process, Process All Traces command.

The processed chromatogram trace can be added to the current Window, or it can replace the
current trace. By default, each subsequent chromatogram, or chromatogram process, is added to
the window; selecting the Tool Bar button causes each subsequent chromatogram, or
chromatogram process, to replace the currently selected trace. Select the button again to toggle it
off.

Note:

The button is grayed when the button is depressed.

Page 6-38
 Chromatogram

Background Subtract
General

Background Subtract fits a smooth curve through the noise in the chromatogram trace, then
subtracts this curve from the chromatogram, leaving the peaks on a flat baseline.

Performing a Background Subtract

To perform Background Subtract:

1. Select the Menu Bar Process, Subtract command; the Background Subtract dialog is
invoked.

2. Select the desired options (see below).

3. Select the OK button; the Background Subtract process starts.

Figure 6.30 The Background Subtract dialog

Polynomial Specifies the degrees of freedom allowed to the fitted curve. With
order polynomial order set to 0, a horizontal straight line is fitted. With
polynomial order set to 1, a sloping straight line is fitted. The further the
background is from a straight line, the higher the Polynomial order value
must be set, however, too high a value will cause the fitted curve to begin
to follow the peak shapes. Normal operating range for this parameter is
3rd to 20th order; the maximum value that can be entered is 99.

Below curve (%) Moves the background curve up and down in the noise. The curve fit is
constrained to place the specified percentage of data points beneath the
fitted background curve. Normal operating range for this parameter is 5%
to 30%, depending on the abundance and width of peaks in the
chromatogram. For fewer or narrower peaks, increase the value. The
maximum value that can be entered is 99%.

Tolerance Affects the precision to which the internal arithmetic is performed. The
permitted range is 0.001 to 0.200; the value should not normally be altered
from its default of 0.010.

Flatten edges Ensures that the polynomial applied is flat (horizontal) at the beginning
and end of the trace.

Make graph of Displays the fitted polynomial itself at the end of the Subtract process,
fitted polynomial rather than the chromatogram with the background curve subtracted.

Page 6-39
Chromatogram

OK Starts the Background Subtract process; the Background Subtract Status

dialog is displayed during processing.

Figure 6.31 The Background Subtract Status dialog

With higher order polynomials, Background Subtract will sometimes have difficulty converging
on a solution. There is a pre-set upper limit of 300 iterations. If Background Subtract does not
seem to be making progress, select the Cancel button in the status box, and try again with a lower-
order polynomial, i.e. with a lower value entered in the Background Subtract dialog Polynomial
order text box.

Example of Background Subtract

The parameters shown in Figure 6.30, when applied to the chromatogram shown in Figure 6.32,
produced the background subtracted chromatogram shown in Figure 6.33.

Figure 6.32 Unprocessed Total Ion Chromatogram

Figure 6.33 Background Subtracted Chromatogram produced using the parameters shown
in Figure 6.30

Page 6-40
 Chromatogram

Checking the Results of Background Subtract

To check the operation of the background subtraction process with a given set of parameters,
select the Background Subtract dialog, Make graph of fitted polynomial option, see the
“Performing a Background Subtract” section, on page 6-39. This causes the same background
subtraction process to take place, but rather than displaying a chromatogram with the background
curve subtracted, the fitted polynomial curve itself is displayed. By choosing Overlay graphs
and Link vertical axes from the Chromatogram Display View dialog (see the “To Change the
Display Parameters” section, on page 6-32), a display like Figure 6.34 can be produced, enabling
the fit of the baseline to the noise to be examined. The parameters shown in Figure 6.30 were
used.

Figure 6.34 Checking the operation of Background Subtract

Smoothing Chromatograms
General

Smoothing improves presentation and aids interpretation of a chromatogram by increasing the

apparent signal-to-noise ratio.

Two types of smoothing are available for chromatograms: Moving Mean and Savitzky Golay.
Both methods slide a window along the chromatogram, averaging the data points in the window to
produce a point in the smoothed chromatogram. Moving Mean takes the arithmetical mean of the
intensities of the data points in the window. Savitzky Golay takes an average of the intensities
weighted by a quadratic curve. This tends to enhance peak and valley shapes, as well as
preserving the height of the peaks better than the Moving Mean. However, Savitzky Golay does
tend to produce small artifacts on either side of the real peaks.

The Smooth Chromatogram Dialog

The Smooth chromatogram dialog is used to control the manner in which smoothing is applied to
a chromatogram; it is invoked by the Menu Bar Process, Smooth command.

Page 6-41
Chromatogram

Figure 6.35 The Smooth chromatogram dialog

Window size Specifies the half-width of the smoothing window, in scans. The
(scans) ± maximum value is 99.

Number of Specifies the number of times the smooth is repeated; increasing this
smooths parameter gives a heavier smooth. The maximum value is 100.

Smoothing
method Frame
Mean Selects the Moving Mean smoothing method.

Savitzky Golay Selects the Savitzky Golay smoothing method.

To Smooth a Chromatogram

1. Select the Menu Bar Process, Smooth command; the Smooth chromatogram dialog is
invoked, see the “The Smooth Chromatogram Dialog” section, on page 6-41.

2. Set the Window size (scans) ± parameter. The number specifies the half-width of the
smoothing window in scans. This parameter can also be set by clicking and dragging across a
chromatogram peak at half height using the right mouse button; the value in the Window size
(scans) ± text box will change to that selected by the mouse, see Figure 6.36.

Figure 6.36 Automatically setting the Window size (scans) ± parameter

3. To alter the number of times the smooth is repeated, change the Number of smooths
parameter from its default value of 2. Increasing this parameter gives a heavier smooth.

4. Select a Smoothing method.

Page 6-42
 Chromatogram

5. Select the OK button; the smoothing process starts. A Smooth message box is displayed
while the calculation is being performed; the smoothed chromatogram is then displayed.

Figure 6.37 Typical Smooth message box

Figure 6.38 Results of chromatogram smoothing

Integrating Chromatograms
General

The Integrate process locates peaks, positions baselines and calculates both the heights and areas
of the peaks above their baselines. There are two possible integration methods available, the
normal algorithm described in the “Standard Peak Detection Parameters” section, on page 6-47,
and the Apex peak integration algorithm, see the “ApexTrack Peak Detection Parameters” section,
on page 6-52.

Both methods can be preceded by smoothing, if this option is chosen. After integration, a
thresholding process can be applied to reject peaks, based on whether their height or area is less
than an absolute value, or less than a specified fraction of the height or area of the largest peak.

There are several stages to the normal integration process:

1. The raw chromatogram is smoothed (if this option is chosen).

2. The data is differentiated with respect to time, and a list of local maxima and minima is
created.

3. The peak-to-peak noise amplitude, measured from the raw chromatogram, defines which
maxima are considered significant. Each local maximum lies between two minima. The
ratios of the intensity (at the maximum value) to the intensity at both neighboring minima are

Page 6-43
Chromatogram

calculated. The maximum represents a peak if both ratios are sufficiently small. Baselines
are initially positioned joining the two neighboring minima. “Shoulder” peaks, that are
completely unresolved, may be detected by searching for local gradient minima on the sides
of each peak.

4. The positions of the baselines are finalized. A parameter may be adjusted to allow the
component peaks of an unresolved multiplet to share a common baseline. Two more
parameters allow compensation for “peak tailing” effects, where the peaks are significantly
asymmetric. The peak baseline will be adjusted to reduce the asymmetry to a specified
maximum.

5. Statistics are calculated for each peak, including peak area by the trapezium rule, and vertical
height of peak top above baseline.

6. Finally the thresholding process is used to reject peaks, based on whether their height or area
is less than an absolute value, or less than a specified fraction of the height or area of the
largest peak.

To Integrate a Chromatogram

A Chromatogram can be integrated, using the current parameters, by selecting the Tool Bar
button. The Integrate parameters can be changed using the Integrate chromatogram dialog; this
is invoked by selecting the Menu Bar Process, Integrate command.

Note:
The integration process operates only on the currently displayed range and not on the whole
chromatogram.

Figure 6.39 The Integrate chromatogram dialog

Noise Frame

Peak-to-peak This value is used to prefilter the chromatogram. A suitable value can be
amplitude measured directly from the chromatogram by clicking the right mouse
button, and dragging the cursor across a section of noise in the
chromatogram, see Figure 6.40. The noise amplitude in this section will
be calculated and the value in the box will be updated. The sensitivity of
the integration algorithm can be fine-tuned by manually adjusting this
value.

Page 6-44
 Chromatogram

Figure 6.40 Setting the peak-to-peak noise amplitude

Automatic Enables automatic measurement of the noise amplitude.

noise
measurement

Enable Performs smoothing (using the current smooth settings) before the
smoothing integration process is begun.

ApexTrack Peak Selects the ApexTrack peak integration algorithm instead of the standard
Integration algorithm for the integration see the “ApexTrack Peak Detection
Parameters” section, on page 6-52, for details.

Note:
The Noise frame is grayed out when this option is selected.

Smooth Invokes the Smooth chromatogram dialog; this is used to control the
manner in which smoothing is applied to a chromatogram, see the “The
Smooth Chromatogram Dialog” section, on page 6-41.

Note:
This button is only available when the Enable smoothing option is
selected.

Peak detect Invokes a dialog that allows the peak detection parameters to be changed.
If the ApexTrack Peak Integration option is not selected, the Peak
Detect dialog is invoked, see the “Standard Peak Detection Parameters”
section, on page 6-47, for details. If the ApexTrack Peak Integration
option is selected, the ApexTrack Peak Detection Parameters dialog is
invoked, see the “ApexTrack Peak Detection Parameters” section, on
page 6-52, for details.

Page 6-45
Chromatogram

Threshold Invokes the Response Threshold dialog; this allows the threshold
parameters (used for optionally removing small peaks) to be changed, see
the “Peak Thresholding” section, on page 6-53, for further details.

Copy Copies the current integration parameters to the Clipboard. These

parameters can then be pasted into another application such as the
Quantify Method Editor.

Paste Pastes a set of integration parameters from the Clipboard.

Note:
This button is grayed out if there are no integration parameters on the
Clipboard.

OK button Exits the dialog and performs the integration; the Peak Integration in
Progress message is displayed. The integration may be stopped at any
time by selecting the Cancel button.

Figure 6.41 Peak Integration in Progress message

Page 6-46
 Chromatogram

Standard Peak Detection Parameters

If the Integrate chromatogram dialog ApexTrack Peak Integration option is not selected (see
the “To Integrate a Chromatogram” section, on page 6-44), the Peak Detect dialog is invoked
when the Peak detect button is selected.

Figure 6.42 The Peak Detect dialog

Page 6-47
Chromatogram

Baselines Frame

Join valleys if Affects how baselines for partially resolved peaks are drawn. The larger
peaks resolved the value of this parameter, the more peak baselines will be drawn up to
to … % above the valleys between unresolved peaks. The default value for this
baseline. parameter is 30%, and the normal operating range is 5% to 75%. The
maximum value is 100%. See Figure 6.43 and Figure 6.44 for examples.

Figure 6.43 Join valleys parameter set to 15%

Figure 6.44 Join valleys parameter set to 60%

Page 6-48
 Chromatogram

Reduce peak Controls the positioning of baseline end points. The default value is 50%,
tailing until and the normal operating range is between 25% and 300%. In the example
trailing edge is below, decreasing the value of the parameter from 150% to 50% reduces
no more the pronounced tail on the peak at 5.42 minutes.
than… %
wider than
leading edge.

Figure 6.45 Reduce peak tailing parameter set to 150%

Figure 6.46 Reduce peak tailing parameter set to 50%

Page 6-49
Chromatogram

Raise baseline Prevents the baseline end point being moved too high up the peak. To
by no more prevent the baseline end points moving up the peaks, reduce the value of
than… % of this parameter. The default value is 10%, and normal operating range is
peak height. 5% to 20%. The maximum value is 100%.

This parameter is only relevant when the Reduce peak tailing parameter
has a small value (less than 50%). (In the example below, the Reduce
peak tailing parameter has been set to 25%.)

Figure 6.47 Raise baseline parameter set to 50%

Figure 6.48 Raise baseline parameter set to 5%

Page 6-50
 Chromatogram

Peak Separation
Frame
Draw vertical Determines how well resolved peaks must be before they are separated by
if peaks a drop line (or baselines are drawn up into the valleys, depending on the
resolved to… value of the Join valleys parameter). Increase the value of this parameter
% above to separate poorly resolved peaks. The default value is 90%, and normal
baseline. operating range is 50% to 100%.

Figure 6.49 Draw verticals parameter set to 50%

Figure 6.50 Draw verticals parameter set to 95%

Detect Select this option to optionally attempt to detect completely unresolved

Shoulder peaks peaks, or shoulders. The algorithm will detect a shoulder if the slope of
if slope is less the shoulder top is less than the specified percentage of the steepest slope
than… % of on the peak. Therefore, to make shoulder detection more sensitive,
maximum. increase the value of this parameter. The default value is 30%, and normal
operating range is 20% to 90%.

OK button Exits the dialog and. returns to the Integrate chromatogram dialog.

Page 6-51
Chromatogram

ApexTrack Peak Detection Parameters

If the Integrate chromatogram dialog ApexTrack Peak Integration option is selected (see the
“To Integrate a Chromatogram” section, on page 6-44), the ApexTrack Peak Detection
Parameters dialog is invoked when the Peak detect button is selected.

Figure 6.51 The ApexTrack Peak Detection Parameters dialog

Peak-to-Peak This is the Apex Detection Threshold. The value of this threshold is the
Baseline Noise maximum (peak-to-peak) excursion of the baseline noise.

ApexTrack converts Peak-to-Peak Baseline Noise to a second derivative

threshold. Peaks that have an inverted second derivative apex higher than
this internal value are considered to be valid peaks. If Automatic is
selected, the second derivative threshold is automatically determined from
the chromatogram. This value is converted to the Peak-to-Peak Baseline
Noise, which is then displayed in the text box; the user-specified value is
ignored. The conversion between Peak-to-Peak Baseline Noise and the
second derivative noise threshold is a factor that depends on the peak
width.

The Peak-to-Peak Baseline Noise is proportional to the second derivative

threshold. Making the Peak-to-Peak Baseline Noise parameter larger will
filter out some of the smaller peaks.

Peak Width at Sets the width, in minutes, of a filter that is used to smooth the second
5% Height derivative. If Automatic is selected, the Peak Width is proportional to the
(Mins) distance between the inflection points (see below) of the highest peak.
The automatic value used is then displayed in the text box and the
user-specified value is ignored. Making this parameter larger will filter
out some of the narrower peaks.

Baseline Start Defines how high the baseline is raised at the start of each peak. It is
Threshold% given as a percentage of the height of the first inflection point of the peak,
i.e. the point on the leading edge of the peak where the second derivative is
zero.

Page 6-52
 Chromatogram

Baseline End Defines how high the baseline is raised at the end of each peak. It is given
Threshold% as a percentage of the height of the last inflection point of the peak, i.e. the
point on the trailing edge of the peak where the second derivative is zero.

If Baseline Start Threshold% and Baseline End Threshold% are both

set to 100%, the baseline will be placed at the peak’s inflection points (see
Figure 6.52).

Detect Shoulders Determines whether any detected shoulders will be treated as separate
peaks or part of the parent peak.

OK button Exits the dialog and. returns to the Integrate chromatogram dialog.

Figure 6.52 Chromatogram Peak and Inverted Second Derivative

Peak Thresholding

Small peaks may be optionally removed by setting one of the four available threshold parameters;
which integrated peak areas and heights are compared against. If a peak’s attribute is less than the
appropriate parameter, the peak is rejected. To examine or edit these parameters, select the
Integrate chromatogram dialog Threshold button (see the “To Integrate a Chromatogram”
section, on page 6-44); the Response Threshold dialog is invoked.

Figure 6.53 The Response Threshold dialog

Page 6-53
Chromatogram

Relative height Removes the peaks whose height is less than the specified percentage of
the highest peak.

Absolute height Removes the peaks whose height is less than the specified value.

Relative area Removes the peaks whose area is less than the specified percentage of the
largest peak area.

Absolute area Removes the peaks whose area is less than the specified value.

OK button Exits the dialog and. returns to the Integrate chromatogram dialog.

To Display Information about an Integrated Peak

Clicking on an integrated peak will display the peak Top position, peak Height and peak Area in
the status bar at the bottom of the Chromatogram Window.

Peak Annotation can be displayed using any combination of peak top time, peak top scan, peak
response height and peak response area by choosing the Menu Bar Display, Peak Annotation
command. This invokes the Chromatogram Peak Annotation dialog, see the “To Change the
Peak Annotation Parameters” section, on page 6-34, for further information.

Editing Integrated Peaks

General

If required, the User can modify integration results by moving the position of an individual
baseline, adding a single peak, or deleting one or more peaks. The Edit Integrated Peaks dialog
(invoked by the Menu Bar Edit, Integrated Peaks command) is used to edit integrated peaks. End
markers appear on the peaks in the chromatogram when the Edit Integrated Peaks dialog has
been invoked.

The Edit Integrated Peaks Dialog

The Edit Integrated Peaks dialog is invoked by the Menu Bar Edit, Integrated Peaks command.

Figure 6.54 The Edit Integrated Peaks dialog

Peak Tops: This is the list of integrated peaks on the current chromatogram trace. A
peak can be selected by clicking on the number in the box. A peak can
also be selected by right-clicking on a peak in the chromatogram trace,
when the Edit Integrated Peaks dialog is on display.

Page 6-54
 Chromatogram

Peak Baseline
Edit Frame
Start The selected peak’s baseline start position; the value can be changed by
typing in a new number.

The start position is also represented on the chromatogram by a small

black square. This can be moved to a new position by clicking and
dragging with the mouse. The value held in the Start box will change to
show the new position.

Any changes in the peak statistics resulting from such modifications are
reflected in the Peak Information frame.

End The selected peak’s baseline end position; the value can be changed by
typing in a new number.

The end position is also represented on the chromatogram by a small black

square. This can be moved to a new position by clicking and dragging
with the mouse. The value held in the End box will change to show the
new position.

Any changes in the peak statistics resulting from such modifications are
reflected in the Peak Information frame.

Add Adds a new baseline having the start and end positions currently specified
in the Start and End boxes. The figures in the Peak Information box are
altered accordingly, and the new peak top is entered into the Peak Tops
box. This button is grayed if the values in the Start and End boxes relate
to an existing, unmodified, peak.

Modify Modifies the start and end positions for an existing peak, as specified in
the Start and End boxes. The figures in the Peak Information box are
altered accordingly. This button is grayed if the values in the Start and
End boxes relate to an existing, unmodified, peak.

Peak Displays information on the currently selected peak.

Information
Frame
Delete Deletes the currently selected Peak Top from the list. The chromatogram
is adjusted by removing the indication of that particular integration from
the trace.

Clear All Deletes all the entries in the Peak Tops box and removes all indications of
integration from the current trace.

OK Saves the changes to the integrated peaks and closes the dialog.

To Edit a Peak Baseline

1. Select the peak whose baseline is to be edited by right-clicking on the peak in the
chromatogram, or by selecting it in the Edit Integrated Peaks dialog Peak Tops: list box.

2. Alter the Start or End point by typing in new values in the Edit Integrated Peaks dialog, or
select a range on the chromatogram with the right mouse button and then select the Edit
Integrated Peaks dialog Modify button.

Page 6-55
Chromatogram

The range can also be changed by clicking on one of the end markers (boxes) on the
chromatogram, and dragging it to the required position.

The figures in the Peak Information frame will update to reflect the edited baseline.

Note:
1. It is possible that a peak’s baseline could be modified in such a way that it would overlap with
another peak’s baseline. In this case the following warning is produced. Select the OK
button, to return the dialog box to the state it was in before the alteration that caused the
error.

Figure 6.55 Peak Edit Error warning: Peak can’t overlap other peaks

2. The Start point must have a lower value than the End point; any attempt to modify a baseline
in contravention to this rule results in the following message. Select the OK button, to return
the dialog box to the state it was in before the alteration that caused the error.

Figure 6.56 Peak Edit Error warning: Start and end positions are invalid

To Add a Peak to the Integration Results

1. Type the start and end points of the new peak’s baseline into the Edit Integrated Peaks
dialog Start and End text boxes, or select a range on the chromatogram with the right mouse
button.

2. Select the Edit Integrated Peaks dialog Add button.

3. The figures in the Peak Information frame will update to reflect the new peak.

To Delete a Peak from the Integration Results

1. Select the peak to be deleted by right-clicking on the peak in the chromatogram, or by

selecting it in the Edit Integrated Peaks dialog Peak Tops: list box.

2. Select the Edit Integrated Peaks dialog Delete button.

To Delete all the Peaks from the Integration Results

Select the Edit Integrated Peaks dialog Clear All button.

Page 6-56
 Chromatogram

Peak Purity
General

The Peak Purity dialog is invoked by the Chromatogram Menu Bar Process, Purity command.

The Peak Purity process works on TIC Chromatograms that have already been integrated.

Note:
It is important not to select the Integrate chromatogram dialog Enable Smoothing option when
integrating the peaks. This is because smoothing tends to increase the peak width, and hence,
when the Purity process selects scans from the edges of the smoothed peak, the scans picked are
actually in the noise in the raw data. Since it is the raw data that is used for the purity
calculation, this will have the effect of artificially depressing the purity value for each peak.

Figure 6.57 The Peak Purity dialog

Method Frame This frame is used to specify which method is to be used for calculating
Peak Purity.

Simple method This method takes no parameters. It works by selecting five spectra from
across the peak, and correlating each spectrum with each other spectrum.
The mean correlation value is displayed, scaled to a percentage (0 to
100%), with 100% representing total purity, and 0% total impurity.

Note:
A purity value of 60% does not mean that the peak has two components in
the ratio 60:40.

Page 6-57
Chromatogram

Bayesian This method requires two parameters; it works by characterizing each

method mass channel as a set of (up to) its first four moments. The first moment
represents peak position, the second peak width, and the third asymmetry.
The program can be restricted to use less than four moments by reducing
the Max no. of moments parameter. Reducing this value will decrease the
runtime of the process. It is also possible to reduce the number of mass
peaks used for comparison. This value is represented by the Max no. of
masses parameter. Decreasing this parameter will also result in reduced
runtime. The Bayesian method is based on a rigorous probabilistic
analysis. The output value loosely represents the natural logarithm of the
probability that the peak is pure. Thus, to calculate the probability that a
peak with purity value x is pure, evaluate exp(x). This implies that the
maximum score (100% probability pure) is zero.

Figure 6.58 Simple Peak Purity

Figure 6.59 Bayesian Peak Purity

Page 6-58
 Chromatogram

To Calculate the Peak Purity Index for a Total Ion Chromatogram

1. Display the chromatogram range of interest in a chromatogram window.

2. Integrate the chromatogram, remembering to disable smoothing.

3. Select the Chromatogram Menu Bar Process, Purity command. The Peak Purity dialog is
invoked.

4. Select the purity method, either Simple or Bayesian.

5. For the Bayesian method, optionally, enter the number of moments to use, and the number of
mass spectral peaks to consider.

6. Select the OK button.

Signal to Noise Ratio

General

It is useful to know the ratio of the peak heights to the level of noise in a mass chromatogram;
MassLynx provides the Signal to Noise process to do this, using the Signal To Noise dialog, see
the “The Signal To Noise dialog” section, on page 6-60.

The Signal to Noise calculations can be performed to display Peak-to-Peak, or RMS values. If
Peak-to-Peak is required, the greatest height of the signal range above the mean noise value is
divided by the span of the noise, where the span of the noise is the difference between the
maximum and minimum values of noise. If RMS is required, the greatest height of the signal
above the mean noise is divided by the root mean square deviation from the mean of the noise.
The RMS is usually expected to be five times the Peak-to-Peak value.

Various authorities have different methods for determining what level of noise is taken into
account for the calculations of noise variance and RMS deviation. A two-step process is carried
out. Firstly, the mean should be calculated with or without zeros as normal. Optional processing
then allows three options:

• The 5% of scans that have the greatest deviation from the mean are disregarded in the noise
signal.

• Those scans whose deviation from the mean is greater than one standard deviation are
disregarded in the noise signal.

• Those scans whose deviation from the mean is greater than two standard deviations are
disregarded in the noise signal.

The first and third options are expected to give roughly equivalent results. The second option
should give an RMS value of about double that of the other two options. If one of these three
processing options is selected (see the “The Signal To Noise dialog” section, on page 6-60) then
the mean and RMS deviation of the noise are recalculated disregarding the appropriate points.

Page 6-59
Chromatogram

Figure 6.60 Signal to Noise processed chromatogram

The Signal To Noise dialog

The Signal To Noise dialog is invoked by selecting the Chromatogram Menu Bar Process, Signal
To Noise command.

Figure 6.61 The Signal to Noise dialog

Ranges Frame

Signal Enter the required range for the signal in this text box, with the start and
end values separated by a colon. Alternatively, right-click and drag across
the required range; the values are automatically entered in the box.

Page 6-60
 Chromatogram

Noise Enter the required range for the noise in this text box, with the start and
end values separated by a colon. Alternatively, right-click and drag across
the required range; the values are automatically entered in the box.

Noise Processing
Frame
Ignore Zeros Ignores zeros.

NO Extra When selected, no extra processing is carried out.

Processing
Ignore Worst The 5% of scans that have the greatest deviation from the mean are
5% of scans disregarded in the noise signal.

Ignore Scans Those scans whose deviation from the mean is greater than one standard
Outside 1 SD deviation are disregarded in the noise signal.

Ignore Scans Those scans whose deviation from the mean is greater than two standard
Outside 2 SD deviations are disregarded in the noise signal.

Display Frame

RMS Displays RMS Signal to Noise values.

Peak to Peak Displays Peak-to-Peak Signal to Noise values.

To Calculate the Signal to Noise Value for a Mass Chromatogram

1. Display the chromatogram range of interest in a chromatogram window.

2. Select the Chromatogram Menu Bar Process, Signal to Noise command; the Signal to Noise
dialog is invoked.

3. Enter the Signal and Noise ranges. Either type values in or select the right mouse button at
one end of the Chromatogram region of interest, and without releasing the button, drag the
cursor horizontally to the other end. As drag the cursor is dragged, a “rubber band” is
stretched out to indicate the range selected. The dialog will be updated to show this range.

4. Select the Noise Processing and Display methods required.

5. Select the OK button.

Combine Spectra

Spectra can be combined, from the Chromatogram window, by selecting the Tool Bar button,
or by selecting the Menu Bar Process, Combine Spectra command; the Combine Spectrum
dialog is invoked. For further details, refer to the “The Combine Spectra Process” section in
Chapter 7, “Spectrum”.

Page 6-61
Chromatogram

ElectroSpray Data Processing – Components

General

In the ElectroSpray spectra of peptides or glycopeptides, that are the result of a digest on an intact
protein or glycoprotein, each component produces a range of multiply-charged ions in the original
m/z spectrum. The range of ions observed depends on the size of the molecule and the number of
charged groups. Most tryptic fragments exhibit at least singly- and doubly-charged ions which
allows unambiguous molecular weight assignment. Small peptides up to 600 Da often only
exhibit a singly-charged ion, but assignment is often possible because of the intensity of the ion in
the spectrum. The molecular mass range of fragments can be anything from 300 to 6000 Da
depending upon digest specificity, i.e. partial cleavages, the type of digest, and whether the
peptides are glycosylated.

Normally, a detailed analysis of a digest and characterization of the resulting peptide fragments
requires several hours of data processing. Auto Find Components combines several processes
(Combine, Mass Measure and Component Finding) to help reduce data processing significantly
and allows the User to accept or reject components visually and interactively.

Component Identification

There are two options available for identifying digest components from an LC/MS analysis:

Either:

Use the Chromatogram Menu Bar Process, Components, Auto Find command to carry out this
operation on a specified range of the LC/MS data file, after Peak Detection (Integration) has been
performed.

Or:

Use the Spectrum Menu Bar Process, Component, Find Auto or Process, Component, Find
Manual command, see Chapter 7, “Spectrum”.

Using either of the above methods will generate the Component Worklist that:

• Provides a summary of components found. Each component is stored in a .cmp file in the raw
data directory.

• Creates a component summary file with extension .cms. This file is stored in the raw data
directory and is used for annotating the chromatogram trace with component labels.

• Interacts with the manual or auto component finding processes in Spectrum. If the Worklist
dialog is active and component finding is carried out from within Spectrum then
Chromatogram and the Worklist dialog are updated to reflect the currently stored component
files. This also applies to editing components from within Spectrum.

Page 6-62
 Chromatogram

The Auto Find Components Dialog

The Auto Find Components dialog is invoked by the Process, Components, Auto Find
command.

Figure 6.62 The Auto Find Components dialog

Combine
parameters
Frame
Combine + -… Refers to the number of scans either side of peak top. If 2 is entered then
scans around five scans in total will be summed around the peak top.
peak top

Component Find
parameters
Frame
Min length… Refers to the minimum number of peaks that form a series of multiply-
peak/s charged ions; e.g. if 2 is entered, this requires that a minimum of two
peaks form a multiply-charged ion series.

Peak Specifies the tolerance on the position of each peak in the series. It may
Window… Da need to be increased from its default value of 0.5 Da for statistically poor
data. Too low a value will result in the algorithm being unable to identify
the whole of the series. Too high a value may result in the algorithm
selecting wrong peaks.

Max std dev… Sets an upper limit on the spread of the molecular masses of the peaks in
Da the series.

% Threshold Specifies a minimum intensity of peaks for the algorithm to consider. It is

specified as a percentage of the intensity of the most intense peak in the
spectrum.

Page 6-63
Chromatogram

Min mol Indicates the lowest molecular mass that the algorithm can consider for a
mass… Da peak series.

Max mol Indicates the highest molecular mass that the algorithm can consider for a
mass…Da peak series.

Identify largest After all the peaks have been associated with a series, this parameter
single peaks specifies how many of the remaining (single) peaks should be associated
with a series.

OK Starts the auto find process; a status box is displayed while processing
takes place.

Mass Measure Invokes the Mass Measure dialog, see the “The Mass Measure Process”
section in Chapter 7, “Spectrum” for details.

Automatic Component Finding

The Auto Find routine finds all components from a peak-detected chromatogram and provides a
summary of all components in a Component Worklist dialog.

1. Peak Detect a selected range of the TIC or BPI traces, see the “Integrating Chromatograms”
section, on page 6-43.

2. Select the Chromatogram Menu Bar Process, Components, Auto Find command. The Auto
Find Components dialog is invoked, see the “The Auto Find Components Dialog” section,
on page 6-63.

3. Enter a Combine + - ….scans around peak top parameter. The default is 2; this means that
two scans either side of the peak top will be used in the combine operation.

4. Enter the Component Find parameters. The Min length should be 2, which requires that a
minimum of two peaks form a multiply-charged ion series. The most important parameter is
the % Threshold which, if set too low, will result in mis-assignments and too many
components for each component file. The Min mol mass value should be twice the lowest
acquired mass and the Max mol mass value should be between 3000 and 4000 for normal
peptides. For a more detailed explanation of these parameters, see the “Finding Components
for Transform” section in Chapter 7, “Spectrum”.

5. If continuum data has been acquired, select the Mass Measure button to set the mass measure
parameters in the Mass Measure dialog, then select the OK button. For more information on
how to use Mass Measure, see the “The Mass Measure Process” section in Chapter 7,
“Spectrum”.

Note:
The Auto Find Components, Mass Measure button is grayed out if centroided data has been
acquired.

6. Select the Auto Find Components dialog box OK button; processing will start.

A status box gives an indication of current processing and allows the operation to be halted by
selecting the Cancel button. Processing time is dependent on the number of peaks detected in the
chromatogram trace, but in most cases, should be complete within 1 or 2 minutes. On completion
of processing, the Component Worklist dialog is displayed showing a summary of all component
files stored on disk. The Spectrum module is also activated to display a multiply-charged
spectrum of the specified combined scan. If continuum data has been acquired, then both the
continuum and centroided data are displayed, see Figure 6.65.

Page 6-64
 Chromatogram

Figure 6.63 Mass Measure dialog parameters

Figure 6.64 Component Worklist and annotated chromatogram

Page 6-65
Chromatogram

Figure 6.65 Combined spectra at scan 129 displaying component assignments. Upper trace
centroided data and lower trace continuum data

The Component Worklist Dialog

General

The Component Worklist dialog is displayed when the Automatic Component Finding process is
completed (see the “Automatic Component Finding” section, on page 6-64). It is also invoked by
the Menu Bar Process, Component, Edit Worklist command.

Note:
There is no direct input into the Worklist. The list of components is read in from a file stored on
disk and displayed in the list box.

Figure 6.66 Component Worklist dialog displaying listing of component files

Main list box Displays the component list.

Secondary list Displays the actual m/z values for the component currently selected in the
box main list box.

Page 6-66
 Chromatogram

Delete Deletes the component currently selected in the main list box.
Components can also be deleted using the keyboard Delete key. This
updates all affected modules in MassLynx.

Sort Sorts components in ascending molecular mass order based on a per

component basis.

Search Copies all highlighted component molecular masses to the Embl database
searching program. This search is termed peptide mapping.

Print Prints the component list.

Copy Copies all highlighted component molecular masses to the BioLynx

program for matching up with theoretical peptide masses that are the result
of a digest.

Match Matches the highlighted components to those in the BioLynx program; see
the “MassLynx NT BioLynx & ProteinLynx Guide” for further
information.

Clear match Clears the selected matches.

Search Seq Searches BioLynx sequences for the selected masses.

While the Component Worklist dialog box is active, any modifications to components from
within the Worklist, or from the Spectrum module, results in the component summary file, .cms,
being updated. This allows Chromatogram, the Worklist and Spectrum to reflect the current status
of stored component files. For example, deleting components from the Worklist or adding new
components from within Spectrum, allows the various windows to be updated and reflect the new
status.

Component labels are assigned in alphabetical order in order of increasing scan number. Labels
continue as AA, AB after Z. The main list box displays the component listing and the secondary
list box displays actual m/z values for each highlighted component. For example, Component A in
Figure 6.66 has a molecular mass of 915.53 and a standard deviation of 0.07. This mass is
calculated from the two peaks at 458.81 (doubly-charged) and 916.47 (singly-charged).

The most important actions in the main list box are:

1. Moving the focus through the list box using the arrow keys on the keyboard or clicking with
the mouse button. This updates the secondary list box.

2. Double-clicking with the mouse, or selecting Enter in the main list box, sends an update
message to Spectrum which is updated to reflect the currently highlighted combined scan.

Selecting and Highlighting Components in the Component Worklist Dialog

Selecting and highlighting components Component Worklist dialog in the main list box is
performed in much the same way as multiple files are selected in Windows Explorer.

More than one component can be selected by holding down the keyboard Ctrl key while clicking
on the components. A block of components can be selected by clicking on the first component in
the block and then holding down the keyboard Shift key while clicking on the last component in
the block. Dragging the mouse cursor down the list performs the same operation. The keyboard
cursor keys may be used instead of the mouse.

Page 6-67
Chromatogram

Deleting Components

Components can be deleted by highlighting them and then selecting the Component Worklist
dialog Delete button, or using the keyboard Delete key. Chromatogram, Spectrum and the
Worklist are updated to reflect the new status of the component files stored on disk.

Sorting Components

Components can be sorted by molecular mass on a per component file basis.

Selecting the Component Worklist dialog Sort button sorts components and updates all the
relevant modules.

Printing Components

A list of components can be obtained in hard-copy format by selecting the Component Worklist
dialog Print button.

Mass Mapping Components

1. Mass mapping can be carried out by searching the component masses against a protein
sequence database. Highlighted component masses can be used in the search. In the example
below all masses were selected from the Component Worklist dialog by dragging the mouse
from the top to the bottom of the list.

2. Selecting the Component Worklist dialog Copy button copies the component masses onto
the clipboard. The Paste button within the ProteinProbe program copies the masses into a
query list (for a more detailed explanation see the “MassLynx NT BioLynx & ProteinLynx
Guide”). The Likelihood scoring scheme is used for ranking hits. It has been demonstrated
that four masses or more are sufficient for uniquely identifying proteins.

Figure 6.67 Mass Mapping component masses against the FASTA protein sequence
database

Page 6-68
 Chromatogram

3. Micromass created indices used in this search. Digest Parameters were set according to
known information about the digest, i.e. tryptic digest, arbitrary mass range of 0 to 200000,
mass error 1 Da and minimum of eight matching masses.

Figure 6.68 Mass Profile Fingerprint identifying beta-lactoglobulin as top likelihood scoring
protein

The top four hits of the search were all beta-lactoglobulins, with the top hit matching nine masses.

Matching Components

Highlighted masses can be matched to a theoretical digest in BioLynx. The component labels for
matched masses changes to that used in BioLynx, e.g. T5, and, if the Chromatogram Peak
Annotation dialog Digest Label option is selected, are also used in annotating the Chromatogram.
The matched components can be unmatched/cleared by selecting the Component Worklist dialog
Clear Match option.

Search Masses against Sequence

Highlighted masses can be searched against a sequence in BioLynx. See the “MassLynx NT
BioLynx & ProteinLynx Guide” for details on output, etc. The BioLynx module has to be running
and active for the search to take place.

Peak Lists
General

The results of peak integration can be saved to disk as a named Peak List file (.pdb). Peak Lists
can then be processed using the MassLynx Quantify program.

The Edit Peak List Dialog

The Edit Peak List dialog is invoked by the Chromatogram Menu Bar Edit, Peak List Write
command. The User can add the results of peak integration to any Peak List. Entries in the Peak
List can be deleted or modified.

Page 6-69
Chromatogram

Figure 6.69 The Edit Peak List dialog

File: Displays the name of the current Peak List file.

Peak Tops: This box lists the current integrated peaks from the active chromatogram
display. A particular peak can be selected by clicking on it in this box, or
by right-clicking on a peak in any visible chromatogram trace.

Peak List: This box lists the peaks associated with the current file. A particular peak
can be selected by clicking on it.

Modify Replaces the selected Peak List peak with the selected Peak Tops peak.
This button is grayed if the Peak List has no entries.

Insert Inserts the selected Peak Tops entry before the selected Peak List entry.

Append Adds the selected Peak Tops entry to the end of the Peak List.

Append All Adds all the Peak Tops entries to the end of the Peak List in the order in
which they appear in the Peak Tops box.

Delete Deletes the selected Peak List entry.

Clear All Deletes all entries in the Peak List.

File Invokes the standard file Open dialog. The supplied defaults are the
current Peak List Drive, Directory and Filename.

A file selected by this dialog becomes the default file for Peak Lists
throughout MassLynx.

Page 6-70
 Chromatogram

To Create a New Peak List File

1. Select the Chromatogram Menu Bar Edit, Peak List Write command. The Edit Peak List
dialog is invoked.

2. Select the File button. The File Open dialog is invoked.

Figure 6.70 The File Open dialog

3. Type the name for the new Peak List file in the File name: text box.

4. Select the Open button. A Create File dialog is invoked.

5. Select the Yes button. The File Open dialog is closed and the Edit Peak List dialog for the
new file appears.

Figure 6.71 The Edit Peak List dialog for a new Peak List file

6. Add peaks to the Peak List: as described in the “To Append Peaks to the Current Peak List”
section, on page 6-72.

7. Select the Exit button. The new Peak List file is saved to disk, the Edit Peak List dialog is
closed, and the new Peak List file becomes the current file.

Page 6-71
Chromatogram

Note:
The new Peak List file is not created if no peaks are added to the Peak List: before selecting the
Exit button.

To Open an Existing a Peak List File

1. Select the Chromatogram Menu Bar Edit, Peak List Write command. The Edit Peak List
dialog is invoked.

2. Select the File button. The File Open dialog is invoked.

3. Select the required file in the list box.

4. Select the Open button. The File Open dialog is closed. The Edit Peak List dialog for the
file appears.

5. Select the Exit button. The Edit Peak List dialog is closed, the selected Peak List file
becomes the current file.

To Append Peaks to the Current Peak List

1. Select the Chromatogram Menu Bar Edit, Peak List Write command. The Edit Peak List
dialog is invoked.

2. Select the peak to be appended either from the Edit Peak List dialog Peak Tops: box, or by
right-clicking on the required peak on a chromatogram trace.

3. Select the Append button.

4. The contents of the Edit Peak List dialog Peak List: box will be updated to include the new
peak.

5. To append all the peaks from Edit Peak List dialog Peak Tops: box, select the Append All
button.

To Delete Peaks from the Current Peak List

1. Select the Chromatogram Menu Bar Edit, Peak List Write command. The Edit Peak List
dialog is invoked.

2. Select the peak to be removed in the Edit Peak List dialog Peak List box.

3. Select the Delete button.

4. To delete all the peaks from Edit Peak List dialog Peak List box, select the Clear All button.

Reading a Peak List into a Chromatogram

The Get Peak List Entry dialog

The Get Peak List Entry dialog is used to select Peak List files; it is invoked by the
Chromatogram Menu Bar Edit, Peak List Read command.

Page 6-72
 Chromatogram

Peak List: Displays the Peak List associated with the current file. Entries may be
selected by clicking on them. Only those peaks that have the same
chromatogram trace are displayed.

Show TIC Displays the Total Ion Current chromatogram.

Figure 6.72 The Get Peak List Entry dialog

Get All Selects the entire Peak List.

If the box is not selected, the chromatogram display will be centered on the
selected Peak List: entry’s stored retention time and the peak’s integration
will be shown.

If this option is selected when the OK button is pressed, the chromatogram

display will refresh to show the whole retention time range with the
selected peaks displayed at their positions in the chromatogram.

OK Selects the current Get All dialog status as the default status. It accepts
the selected peaks as those to be displayed. If it is not possible to create a
Chromatogram using this information the warning message box appears
and no further processing is performed.

File Invokes the standard file Open dialog. The supplied defaults are the
current Peak List Drive, Directory and Filename. A file selected by this
dialog becomes the default file for Peak Lists throughout MassLynx.

To Select a Peak List File

1. Select the Chromatogram Menu Bar Edit, Peak List Read command; the Get Peak List
Entry dialog for the current file is invoked.

2. Select the File button; the File Open dialog is invoked.

3. Select the required file in the list box.

4. Select the Open button. The File Open dialog is closed and the Get Peak List Entry dialog
for the selected file appears.

5. Select the OK button.

Page 6-73
Chromatogram

To Read a Single Peak into the Currently Selected Chromatogram

1. Select the Chromatogram Menu Bar Edit, Peak List Read command. The Get Peak List
Entry dialog for the current file is invoked.

2. Select a peak by clicking in the Peak List: box.

3. Select the OK button.

To Read a Whole Peak List into the Currently Selected Chromatogram

1. Select the Chromatogram Menu Bar Edit, Peak List Read command. The Get Peak List
Entry dialog for the current file is invoked.

2. Select the Get All option.

3. Select the OK button.

Chromatogram Display When Switching Between

MS and MS/MS Modes
When an instrument is being used for Data Dependant Acquisition (DDA) and parent ion
discovery (parent and neutral loss scanning), the chromatogram drops to zero when the instrument
is in MS mode. This makes it easier to see when the mass spectrometer has switched between the
MS/MS and MS functions.

The upper trace in Figure 6.73 shows a typical MS/MS TIC chromatogram dropping to zero when
the instrument is in MS mode. The retention times and set masses are annotated on this
chromatogram. The lower trace shows the corresponding MS TIC chromatogram.
BCASmix2 3: TOF MSMS ES+
20.45 TIC
100
577.27 1.04e4

19.29
1031.43

%
18.47
542.22

16.58
432.20 20.78 27.91
23.51 24.65 26.65
765.35 22.12 669.29
533.23 960.44 570.27
590.29
0
BCASmix2 1: TOF MS ES+
18.79 TIC
100
2.55e5
19.20

27.61
20.27 20.98
21.39

23.71 26.02

% 26.47
22.44 23.08 25.38
16.70
15.19 16.09
28.81

0 Time
14.00 15.00 16.00 17.00 18.00 19.00 20.00 21.00 22.00 23.00 24.00 25.00 26.00 27.00 28.00 29.00

Figure 6.73 MS/MS TIC chromatogram dropping to zero when the instrument is in MS
mode

Page 6-74
 Chromatogram

Copying To and From the Windows Clipboard

General

The Windows Clipboard can be used to move data into or out of the Chromatogram window,
either as a picture, or as a text list. For example, spectra or chromatograms can be pasted into
reports written with a Windows compatible word processor.

To Copy a Chromatogram as a Picture to the Clipboard

1. Produce the required display in the Chromatogram window.

2. Select the Tool Bar button, or select the Chromatogram Menu Bar Edit, Copy Picture
command. The contents of the window are copied to the Clipboard as both a metafile and a
bitmap.

To Copy a Chromatogram as a Text List to the Clipboard

1. Display the required time range in the Chromatogram window.

2. Select the Tool Bar button, or select the Chromatogram Menu Bar Edit, Copy
Chromatogram List command. The displayed section of the chromatogram will be copied to
the Clipboard as (time, intensity) pairs or (scan, intensity) pairs depending on the horizontal
axis setting.

To Copy Integrated Chromatogram Peaks as a Text List to the

Clipboard

1. Display the required time range in the Chromatogram window.

2. Select the Tool Bar button, or select the Chromatogram Menu Bar Edit, Copy Detected
Peaks command. The displayed chromatogram peaks will be copied to the Clipboard. The
information transferred for each peak is the peak top, height, area, start, end, start height and
end height.

To Paste Information from the Windows Clipboard into a

Chromatogram Window

1. Select the Tool Bar button, or select the Chromatogram Menu Bar Edit, Paste command
to paste the default Clipboard object to chromatogram. Select the Edit, Paste Special
command to select which object to paste into the Chromatogram. These objects would
typically be metafiles, bitmaps, or text.

2. Use the mouse to drag the outline of the image to the required position.

Any contents of the Clipboard, be it a bitmap, a metafile or text, can be pasted into a
chromatogram window. If the data is in textual or metafile form, it can be re-scaled using the
mouse, and there will be no distortion of the image. However, if a bitmap is pasted, re-scaling is
done by stretching the image; this will cause some distortion. To avoid this, scale the image to the
required size before copying it to the Clipboard.

Page 6-75
Chromatogram

Removing Pasted Input from the Display

1. Click on the item to be removed.

2. Press the keyboard Delete key.

Retention Index
General

The Retention Index is used to compare results from different HPLC systems and different
columns. LogP is a measure of the hydrophobicity.

The Retention Index Table dialog

The Retention Index Table dialog is invoked by the Menu Bar Tools, Retention Index, Edit
Index Table command.

Figure 6.74 The Retention Index Table dialog

Retention Index A set of values is provided with standard compounds. These values are
entered in this text box. Click on an entry in the list of retention indices to
display its Retention Index value in this box.

LogP A set of values is provided with standard compounds. These values are
entered in this text box. Click on an entry in the list of retention indices to
display its LogP value in this box.

Add Adds the Retention Index and LogP values to the bottom of the list of
retention indices.

Page 6-76
 Chromatogram

Sort Sorts the table in order of increasing retention index.

Modify To modify an entry, click on the entry in the list, change the values in the
Retention Index and LogP boxes, and select the Modify button.

Delete Deletes the currently-selected entry from the list.

To Set Up the Retention Index Table

1. Select the Chromatogram Menu Bar Retention Index, Edit Index Table command. The
Retention Index Table dialog is invoked.

2. A set of values will be supplied with the standard compound; enter these values in the table.

To add an entry, type in a Retention Index and LogP value supplied with the standard
compound, and select the Add button.

To modify an entry, click on the entry in the list, change the values in the Retention Index
and LogP boxes, and select the Modify button.

To delete an entry, click on the entry in the list, and select the Delete button.

Selecting the Sort button sorts the list in order of ascending Retention Index.

3. Run the standard compound to assign real times to the Retention Index values.

To Delete the Retention Index Table

1. Select the Chromatogram Menu Bar Tools, Retention Index, Delete Index Table command.
The User is prompted to confirm the deletion.

2. Select Yes to delete the Retention Index Table.

To Make a Retention Index Calibration

1. Integrate the chromatogram; ensure that smoothing is disabled.

2. Select the Chromatogram Menu Bar Tools, Retention Index, Make Calibration command.
The Make Retention Index Calibration dialog is invoked.

Figure 6.75 The Make Retention Index Calibration dialog

Page 6-77
Chromatogram

Start index in To calibrate over the same range as the standard, set this value to 1. To
table calibrate over a different range enter the number of the entry in the
Retention Index Table dialog at which to start.

End index in To calibrate over the same range as the standard, set this to the number of
table the last entry in the Retention Index Table dialog. To calibrate over a
different range enter the number of the entry in the Retention Index Table
dialog at which to end.

Peak Difference This is normally set to 1 to measure all the peaks. If small secondary
peaks appear, the Peak Difference can be set to a higher number so that the
secondary peaks are not used in the calibration.

When a Retention Index calibration is performed, MassLynx matches peaks in the trace with those
in the Retention Index Table and assigns a real time to the Retention Index value. MassLynx then
interpolates the results and displays Retention Index values for each peak in the chromatogram
trace.

Figure 6.76 Retention Index calibration curve

To Display Retention Index Values on a Chromatogram

1. Select the Menu Bar Display, View command. The Chromatogram Display View dialog is
invoked, see Figure 6.25, on page 6-32.

2. Select the Peak List option.

3. Select the OK button.

Page 6-78
 Chromatogram

To Check Retention Index Calibration Status

Select the Chromatogram Menu Bar Tools, Retention Index, Calibration Status command.

If calibration has been performed then the

dialog will be displayed, otherwise, the

dialog will be displayed.

Page 6-79
Chromatogram

Page 6-80
 Spectrum

Chapter 7 Spectrum

Page 7-1
Spectrum

Contents
Getting Started............................................................................................................................... 7-7
To Display the First Scan of the Current Data File ............................................................ 7-7
To Display a Scan at a Particular Time in the Current Data File ....................................... 7-7
The Spectrum Window.................................................................................................................. 7-8
The Spectrum Menu Bar ............................................................................................................... 7-9
The Spectrum File Menu.................................................................................................... 7-9
The Spectrum Edit Menu ................................................................................................... 7-9
The Spectrum Display Menu............................................................................................ 7-10
The Spectrum Process Menu............................................................................................ 7-13
The Spectrum Tools Menu............................................................................................... 7-15
The Spectrum Window Menu .......................................................................................... 7-16
The Spectrum Help Menu ................................................................................................ 7-17
The Spectrum Tool Bar ............................................................................................................... 7-17
General............................................................................................................................. 7-17
Customizing the Spectrum Tool Bar ................................................................................ 7-18
Displaying Spectra ...................................................................................................................... 7-21
Adding or Replacing Spectra ........................................................................................... 7-21
The New Spectrum Dialog............................................................................................... 7-22
Viewing a Peak List Entry ............................................................................................... 7-22
Manipulating the Display ............................................................................................................ 7-23
Altering the Range of the Mass Axis ............................................................................... 7-23
Altering the Range of the Intensity Axis.......................................................................... 7-23
Altering the Range of Both Axes ..................................................................................... 7-23
Setting Magnified Ranges ................................................................................................ 7-23
Deleting Magnification Ranges........................................................................................ 7-25
Restoring the Display....................................................................................................... 7-25
Setting the Display Range Defaults ................................................................................. 7-26
Displaying a Spectrum as a List....................................................................................... 7-26
Controlling the Appearance of the Display ................................................................................. 7-28
General............................................................................................................................. 7-28
To Change the Display Parameters .................................................................................. 7-28
Controlling the Appearance of Peak Labels ................................................................................ 7-31
General............................................................................................................................. 7-31
To Change the Peak Annotation Parameters .................................................................... 7-31
To Annotate a Particular Peak.......................................................................................... 7-33
Removing Spectra from the Display ........................................................................................... 7-33
To Remove a Single Spectrum Trace from the Display................................................... 7-33
To Remove Multiple Spectrum Traces from the Display................................................. 7-33
Real-Time Display of Spectra .......................................................................................... 7-33
Changing the Order of Displayed Spectra........................................................................ 7-34
Adding Text to the Spectrum Display.............................................................................. 7-34
Exporting SEQUEST Files.......................................................................................................... 7-34
General............................................................................................................................. 7-34
To Export a SEQUEST File ............................................................................................. 7-35
Processing Spectra....................................................................................................................... 7-35
General............................................................................................................................. 7-35
Saving and Recalling Processed Spectra.......................................................................... 7-36
The Refine Process...................................................................................................................... 7-38
General............................................................................................................................. 7-38
To Refine a Scan in a Centroid-Mode Data File .............................................................. 7-38
The Combine Spectra Process ..................................................................................................... 7-39
General............................................................................................................................. 7-39
To Combine Scans in a Centroid-Mode Data File ........................................................... 7-39
The Background Subtract Process............................................................................................... 7-40
General............................................................................................................................. 7-40
To Subtract the Background from a Continuum Spectrum .............................................. 7-40
The Smooth Process .................................................................................................................... 7-41

Page 7-2
 Spectrum

General............................................................................................................................. 7-41
To Smooth a Continuum Spectrum.................................................................................. 7-42
The Center Process...................................................................................................................... 7-43
General............................................................................................................................. 7-43
To Center a Continuum Spectrum ................................................................................... 7-44
The Mass Measure Process ......................................................................................................... 7-46
General............................................................................................................................. 7-46
QTOF Accurate Mass ...................................................................................................... 7-47
The TOF Transform Process ....................................................................................................... 7-47
The Integration Process............................................................................................................... 7-48
General............................................................................................................................. 7-48
To Integrate a Spectrum................................................................................................... 7-48
ElectroSpray Data Processing ..................................................................................................... 7-49
General............................................................................................................................. 7-49
Setting Adduct Mass for Transform and MaxEnt ............................................................ 7-50
Finding Components for Transform................................................................................. 7-50
Editing Components for Transform ................................................................................. 7-54
The Transform Process .................................................................................................... 7-58
MaxEnt 1 ......................................................................................................................... 7-59
MaxEnt Errors.................................................................................................................. 7-68
MaxEnt 2 ......................................................................................................................... 7-69
MaxEnt 3 ......................................................................................................................... 7-71
Isotope Cluster Abundance Plots ................................................................................................ 7-75
General............................................................................................................................. 7-75
The Isotope modelling Dialog ......................................................................................... 7-75
The User-definable elements Dialog................................................................................ 7-77
To Produce an Isotope Cluster Abundance Plot............................................................... 7-78
Elemental Composition ............................................................................................................... 7-78
General............................................................................................................................. 7-78
The EleComp Parameters Dialog..................................................................................... 7-78
To Produce an Elemental Composition Report................................................................ 7-79
To Update an Elemental Composition Report ................................................................. 7-80
The Elemental Composition Window.............................................................................. 7-80
The Elemental Composition Window Menu Bar............................................................. 7-82
The Elemental Composition Window Tool Bar............................................................... 7-84
Elemental Composition Parameters ............................................................................................ 7-84
General............................................................................................................................. 7-84
The Parameters Dialog; General Parameters Page........................................................... 7-85
The Parameters Dialog; Symbol Parameters Page........................................................... 7-87
Superatom Tables ............................................................................................................ 7-91
Superatom Limits............................................................................................................. 7-93
Performing a Calibration............................................................................................................. 7-94
General............................................................................................................................. 7-94
To Make a New Calibration............................................................................................. 7-94
To Apply a Calibration .................................................................................................... 7-96
To Modify a Calibration .................................................................................................. 7-97
Lock Mass........................................................................................................................ 7-97
Copying to and from the Windows Clipboard ............................................................................ 7-97
General............................................................................................................................. 7-97
To Copy a Spectrum as a Picture to the Clipboard .......................................................... 7-98
To Copy a Spectrum as a Text List to the Clipboard ....................................................... 7-98
To Paste Information from the Windows Clipboard into a Spectrum Window ............... 7-98
Removing Pasted Input from the Display ........................................................................ 7-98
Manipulating Library Spectra ..................................................................................................... 7-99
To Display a Library Entry .............................................................................................. 7-99
To Append the Current Spectrum to the Current Library ................................................ 7-99

Page 7-3
Spectrum

Illustrations
Figure 7.1 The Select Raw Spectrum dialog ................................................................................ 7-7
Figure 7.2 The Display Raw Spectrum dialog.............................................................................. 7-7
Figure 7.3 The Spectrum Window ............................................................................................... 7-8
Figure 7.4 The Spectrum File Menu............................................................................................. 7-9
Figure 7.5 The Spectrum Edit Menu ............................................................................................ 7-9
Figure 7.6 The Edit, Library sub-menu ...................................................................................... 7-10
Figure 7.7 The Spectrum Display menu ..................................................................................... 7-10
Figure 7.8 The Spectrum Display, Spectrum sub-menu............................................................. 7-11
Figure 7.9 The Spectrum Display, Range sub-menu .................................................................. 7-12
Figure 7.10 The Spectrum Process Menu................................................................................... 7-13
Figure 7.11 The Spectrum Process, Component sub-menu........................................................ 7-15
Figure 7.12 The Spectrum Tools Menu...................................................................................... 7-15
Figure 7.13 The Spectrum Window Menu ................................................................................. 7-16
Figure 7.14 The Customize Toolbar dialog................................................................................ 7-19
Figure 7.15 The New Spectrum dialog....................................................................................... 7-22
Figure 7.16 The Display Quan DB Spectrum dialog.................................................................. 7-22
Figure 7.17 The Display Range dialog....................................................................................... 7-23
Figure 7.18 The Spectrum Magnify dialog................................................................................. 7-24
Figure 7.19 The Default Spectrum Range dialog ....................................................................... 7-26
Figure 7.20 Typical spectrum displayed as a list........................................................................ 7-27
Figure 7.21 The Spectrum Print Report dialog........................................................................... 7-27
Figure 7.22 The Spectrum Display dialog.................................................................................. 7-28
Figure 7.23 The Spectrum Peak Annotation dialog.................................................................... 7-31
Figure 7.24 The Short Cut Remove Spectrum dialog................................................................. 7-33
Figure 7.25 The Remove Spectra dialog .................................................................................... 7-33
Figure 7.26 The Spectrum Real-Time Update dialog................................................................. 7-34
Figure 7.27 The Export SEQUEST compatible file dialog ........................................................ 7-35
Figure 7.28 The Spectrum Save dialog ...................................................................................... 7-36
Figure 7.29 The Spectrum Data Browser dialog ........................................................................ 7-37
Figure 7.30 The History Selector dialog .................................................................................... 7-37
Figure 7.31 The Refine Spectrum dialog.................................................................................... 7-38
Figure 7.32 The Combine Spectrum dialog................................................................................ 7-39
Figure 7.33 The Background Subtract dialog............................................................................. 7-41
Figure 7.34 The Spectrum Smooth dialog.................................................................................. 7-42
Figure 7.35 The Spectrum Center dialog.................................................................................... 7-44
Figure 7.36 The Mass measure dialog........................................................................................ 7-46
Figure 7.37 The TOF Accurate Mass dialog .............................................................................. 7-47
Figure 7.38 The TOF Transform dialog ..................................................................................... 7-48
Figure 7.39 The Peak Detect dialog ........................................................................................... 7-49
Figure 7.40 The Set Adduct Mass dialog ................................................................................... 7-50
Figure 7.41 The Manual Find Components dialog ..................................................................... 7-51
Figure 7.42 The Automatic Find Components dialog ................................................................ 7-52
Figure 7.43 The Edit Components dialog................................................................................... 7-55
Figure 7.44 The Edit Component dialog .................................................................................... 7-56
Figure 7.45 The Transform dialog.............................................................................................. 7-58
Figure 7.46 The MaxEnt 1 dialog............................................................................................... 7-60
Figure 7.47 Theoretical Peak Width of Proteins due to Isotopic Distribution
vs. Molecular Weight .............................................................................................. 7-64
Figure 7.48 First three iterations of a MaxEnt survey run on leech haemoglobin...................... 7-65
Figure 7.49 Original data from leech haemoglobin (lower), and MaxEnt mock data (upper).... 7-66
Figure 7.50 MaxEnt results from leech haemoglobin................................................................. 7-67
Figure 7.51 The MaxEnt 2 dialog............................................................................................... 7-69
Figure 7.52 The MaxEnt Reconstruction status dialog............................................................... 7-70
Figure 7.53 Original data from Peptide mixture (upper and middle) and
MaxEnt 2 data (lower) ............................................................................................ 7-71
Figure 7.54 The MaxEnt 3 dialog (TOF data)............................................................................ 7-72
Figure 7.55 The MaxEnt 3 dialog (Quad data)........................................................................... 7-73
Figure 7.56 The MaxEnt 3 Advanced Parameters dialog ........................................................... 7-73

Page 7-4
 Spectrum

Figure 7.57 The MaxEnt3 Sequence status dialog ..................................................................... 7-74

Figure 7.58 Original data from Glu-fibrinopeptide (lower) and MaxEnt 3 data (upper)............ 7-74
Figure 7.59 Typical Isotope modeling display ........................................................................... 7-75
Figure 7.60 The Isotope modelling dialog ................................................................................. 7-76
Figure 7.61 The User-definable elements dialog ....................................................................... 7-77
Figure 7.62 The EleComp Parameters dialog............................................................................. 7-78
Figure 7.63 The Mass dialog...................................................................................................... 7-79
Figure 7.64 The Isotope Cluster Parameters dialog ................................................................... 7-80
Figure 7.65 Typical Elemental Composition Window............................................................... 7-80
Figure 7.66 The Elemental Composition Window File Menu ................................................... 7-82
Figure 7.67 The Elemental Composition Window Edit Menu ................................................... 7-82
Figure 7.68 The Elemental Composition Window View Menu ................................................. 7-83
Figure 7.69 The Elemental Composition Window Process Menu ............................................. 7-83
Figure 7.70 The Elemental Composition Window Help Menu.................................................. 7-83
Figure 7.71 The Parameters dialog: General Parameters page................................................... 7-85
Figure 7.72 The Parameters dialog: Symbol Parameters page ................................................... 7-87
Figure 7.73 The Select Hot Symbol dialog ................................................................................ 7-88
Figure 7.74 The Periodic Table dialog....................................................................................... 7-89
Figure 7.75 The Isotopes dialog ................................................................................................. 7-90
Figure 7.76 The Superatom Tables dialog ................................................................................. 7-92
Figure 7.77 The Superatom Limits dialog.................................................................................. 7-93
Figure 7.78 The Elemental Composition warning box............................................................... 7-93
Figure 7.79 The Make new calibration dialog............................................................................ 7-95
Figure 7.80 The Calibration Report window.............................................................................. 7-95
Figure 7.81 The Calibration Parameters dialog.......................................................................... 7-96
Figure 7.82 The Apply Calibration dialog ................................................................................. 7-96
Figure 7.83 The Modify Calibration dialog ............................................................................... 7-97
Figure 7.84 The Lock Mass dialog............................................................................................. 7-97
Figure 7.85 The Display Library Spectrum dialog..................................................................... 7-99
Figure 7.86 The Append Spectrum dialog ................................................................................. 7-99

Page 7-5
Spectrum

Page 7-6
 Spectrum

Getting Started
To Display the First Scan of the Current Data File

Select the MassLynx Sample List Menu Bar Spectrum command, the first scan of the current data
file is displayed.

To Display a Scan at a Particular Time in the Current Data File

Either:

Double-click at the required time (on the X-axis) in the Chromatogram display. The
Spectrum for that time is displayed.

Or:

Select the Spectrum Tool Bar button. The Select Raw Spectrum dialog is invoked; the
Entry: box displays the time for the currently-displayed spectrum.

Figure 7.1 The Select Raw Spectrum dialog

a. Enter the time for the required Spectrum in the Entry: text box.

b. Select the OK button. The Spectrum for that time is displayed.

Or:

Select the Spectrum Menu Bar Display, Spectrum, At command. The Display Raw
Spectrum dialog is invoked; the Spectrum: text box displays the time for the currently
displayed spectrum.

Figure 7.2 The Display Raw Spectrum dialog

a. Enter the time for the required Spectrum in the Spectrum: text box.

b. Select the OK button. The Spectrum for that time is displayed.

Page 7-7
Spectrum

The Spectrum Window

Figure 7.3 The Spectrum Window

The Spectrum module runs in a top-level window that has a Menu Bar and Tool Bar at the top.

The top-level window may contain one or more Spectrum Windows; each can contain one or more
Spectrum traces.

When there is more than one trace in a window, the current trace is identified by a colored square
at the left of the trace. To select another trace, click on any part of the trace, or select a trace from
the Menu Bar Display, Graphs command, or use the keyboard up and down arrow keys.

The spectra in each Spectrum window share a common mass axis; place Spectra in separate
windows to display them on different mass axes.

Page 7-8
 Spectrum

The Spectrum Menu Bar

The Spectrum File Menu

Figure 7.4 The Spectrum File Menu

Open Opens a data file.

Save Spectrum Saves a processed spectrum, see the “To Save a Processed Spectrum”
section, on page 7-36.

Note:
This option is only available when a processed spectrum is selected.

Export Allows the export of data to an ASCII file consisting of spectrum masses
SEQUEST file and intensities, see the “Exporting SEQUEST Files” section, on page 7-34.

Note:
This option is only available for BioLynx and non-GC installations.

Print Prints the current Spectrum Window.

Print Report Prints a list of spectrum masses and intensities, see the “To Print a Report
of the Spectrum Listing” section, on page 7-27.

Printer Setup Invokes the standard Windows Print Setup dialog.

Exit Closes the Chromatogram window.

The Spectrum Edit Menu

Figure 7.5 The Spectrum Edit Menu

Copy Picture Copies the current Window to the clipboard.

Page 7-9
Spectrum

Copy Spectrum Copies the currently-displayed range of the spectrum trace to the clipboard
List as mass and intensity pairs in the form of a text list.

Paste Pastes the clipboard contents into the display.

Paste Special Invokes the standard Windows Paste Special dialog.

Library Invokes the Library sub-menu, used to append spectra to user libraries and
to view spectra in any library.

Get Spectrum Invokes the Display Library Spectrum dialog, which allows a Library
entry to be displayed, see the “To Display a Library Entry” section, on
page 7-99.

Append Invokes the Append Spectrum dialog, which is used to append the
current spectrum to the Library, see the “To Append the Current Spectrum
to the Current Library” section, on page 7-99.

Figure 7.6 The Edit, Library sub-menu

The Spectrum Display Menu

Figure 7.7 The Spectrum Display menu

Spectrum Invokes the Spectrum sub-menu, see the “The Spectrum Display,
Spectrum Sub-Menu” section, on page 7-11.

Remove Invokes the Remove Spectra dialog, used to remove multiple spectrum
traces from the display; see the “To Remove Multiple Spectrum Traces
from the Display” section, on page 7-33.

Page 7-10
 Spectrum

Real-Time Invokes the Spectrum Real-Time Update dialog, used to display new
Update spectra as they are being acquired, see the “Real-Time Display of Spectra”
section, on page 7-33.

Range Invokes the Range sub-menu, see the “The Spectrum Display, Range Sub-
Menu” section, on page 7-12 for details.

List Spectrum Displays a spectrum as a list of peak masses and intensities, see the
“Displaying a Spectrum as a List” section, on page 7-26.

View Invokes the Spectrum Display dialog, used to change the spectrum
display parameters, see the “Controlling the Appearance of the Display”
section, on page 7-28.

Peak Annotation Invokes the Spectrum Peak Annotation dialog, used to edit the Peak
Annotation Parameters; see the “Controlling the Appearance of Peak
Labels” section, on page 7-31.

Customize Invokes the Customize Toolbar dialog; see the “Customizing the
Toolbar Spectrum Tool Bar” section, on page 7-18.

Toolbar Toggles the Tool Bar on and off.

Status bar Toggles the Status Bar on and off.

Move to Last Moves the currently selected Spectrum to the top of the display, see the
“Changing the Order of Displayed Spectra” section, on page 7-34 for
further details.

Move to First Moves the currently selected Spectrum to the bottom of the display, see
the “Changing the Order of Displayed Spectra” section, on page 7-34 for
further details.

Graphs Displays a list of the Spectra in the display; click on a Spectrum in the list
to select it.

The Spectrum Display, Spectrum Sub-Menu

Figure 7.8 The Spectrum Display, Spectrum sub-menu

Page 7-11
Spectrum

At Invokes the Display Raw Spectrum dialog, this allows the User to view a
spectrum, see the “To Display a Scan at a Particular Time in the Current
Data File” section, on page 7-7.

Peak List Invokes the Display Quan DB Spectrum dialog, this allows the User to
Entry view a Peak List entry, see the “Viewing a Peak List Entry” section, on
page 7-22.

The Spectrum Display, Range Sub-Menu

Figure 7.9 The Spectrum Display, Range sub-menu

From Invokes the Display Range dialog, used to change the mass axis range;
see the “To Alter the Range of the Mass Axis using the Menu Bar”
section, on page 7-23.

Magnify Invokes the Spectrum Magnify dialog, used to magnify a section of the
current spectrum trace; see the “Setting Magnification Ranges using the
Menu Bar Magnify Command” section, on page 7-24.

Default Invokes the Default Spectrum Range dialog, to specify the default mass
axis range; see the “To Change the Default Display Range” section, on
page 7-26.

Page 7-12
 Spectrum

The Spectrum Process Menu

Note:
Spectrum only enables those processes that can be applied to the currently loaded data; hence,
menu commands that are not applicable to the data are grayed-out.

Figure 7.10 The Spectrum Process Menu

Refine Invokes the Refine Spectrum dialog, used to automatically remove

background ions from a spectrum thereby allowing it to be more easily
identified, for example by library search, see the “The Refine Process”
section, on page 7-38.

Note:
The Refine process operates on centroid-mode data only.

Combine Invokes the Combine Spectrum dialog, used to produce a single spectrum
by subtracting averaged background spectra from the average of spectra
from a TIC peak, see the “The Combine Spectra Process” section, on
page 7-39.

Subtract Invokes the Background Subtract dialog, used to adjust the zero level in
a continuum spectrum to lessen the effect of chemical noise caused by
column bleed, etc, see the “The Background Subtract Process” section, on
page 7-40.

Smooth Invokes the Spectrum Smooth dialog, used to reduce the high-frequency
noise present in a spectrum, thus aiding interpretation, see the “The
Smooth Process” section, on page 7-41.

Page 7-13
Spectrum

Center Invokes the Spectrum Center dialog, used to calculate the mass of the
peak center, see the “The Center Process” section, on page 7-43.

Note:
This option is disabled for non-continuum data.

Mass Measure Invokes the Mass Measure dialog, used to center peaks with optional
background subtraction and/or smoothing, on continuum spectra, see the
“The Mass Measure Process” section, on page 7-46.

Process All Select once to process all traces in the current window. Select again to
Traces process only the current trace in the current window.

Component Invokes the Component sub-menu, see the “The Spectrum Process,
Component Sub-Menu” section, on page 7-15.

Note:
This Component sub-menu is only available for ElectroSpray data.
ElectroSpray data consists of a series of multiply-charged ions. This
series identifies a component that is used in the Transform and MaxEnt
processes.

Transform Invokes the Transform dialog, see the “To Transform an ElectroSpray
Spectrum onto a Molecular Mass Axis; The Transform dialog” section, on
page 7-58.

MaxEnt 1 Invokes the MaxEnt 1 dialog, used to produce true molecular mass spectra
from multiply-charged ElectroSpray spectra, see the “MaxEnt 1” section,
on page 7-59.

MaxEnt Errors Calculates a probable error range for the mass of each peak in the MaxEnt
spectrum, see the “MaxEnt Errors” section, on page 7-68.

Set Adduct Mass Invokes the Set Adduct Mass dialog used to set the adduct mass for
MaxEnt and Transform, see the “Setting Adduct Mass for Transform and
MaxEnt” section, on page 7-50.

MaxEnt 2 Invokes the MaxEnt 2 dialog, used to increase resolution and remove
noise for any singly charged continuum spectrum, see the “MaxEnt 2”
section, on page 7-69.

MaxEnt 3 Invokes the MaxEnt 3 dialog, used to resolve the multiply-charged peaks
onto a singly-charged axis for any low mass, multiply-charged continuum
spectrum, see the “MaxEnt 3” section, on page 7-71.

Integrate Invokes the Peak Detect dialog, used to locate spectral peaks, draw
baselines and calculate peak areas, see the “To Integrate a Spectrum”
section, on page 7-48.

TOF Transform Invokes the TOF Transform dialog, see the “The TOF Transform
Process” section, on page 7-47.

Page 7-14
 Spectrum

The Spectrum Process, Component Sub-Menu

Figure 7.11 The Spectrum Process, Component sub-menu

Edit Invokes the Edit Components dialog, used to edit the components stored
in the component table, see the “Editing Components for Transform”
section, on page 7-54.

Find Auto Invokes the Automatic Find Components dialog, used to automatically
find components when the mass range is known, see the “To Find
Components when the Mass Range is Known; the Automatic Find
Components Dialog” section, on page 7-52.

Find Manual Invokes the Manual Find Components dialog, used to manually find
components when the mass range is unknown, see the “To Find
Components using the Manual Method; the Manual Find Components
Dialog” section, on page 7-51.

The Spectrum Tools Menu

Figure 7.12 The Spectrum Tools Menu

Page 7-15
Spectrum

Library Search Invokes the Library Search List dialog, used to identify the current scan
using the library search facility, see Chapter 11 “Library”.

Note:
This option is only enabled for centroid data.

Isotope Model Invokes the Isotope modelling dialog, used to produce an isotope cluster
abundance plot for a given formula, see the “The Isotope modelling
Dialog” section, on page 7-75.

Elemental Invokes the EleComp Parameters dialog, used to search for the possible
Composition component element(s) of a selected peak, see the “The EleComp
Parameters Dialog” section, on page 7-78.

ACD Labs Invokes the Advanced Chemistry Development's ACD/Spec Manager

SpecManager software suite for structural elucidation, (if installed on the PC).

Make Invokes the Make new calibration dialog, used to make a calibration
Calibration using a reference file, see the “Performing a Calibration” section, on
page 7-94.

Apply Invokes the Apply Calibration dialog, used to apply the calibration
Calibration previously made using the Make Calibration command, see the “To
Apply a Calibration” section, on page 7-96

Modify Invokes the Modify Calibration dialog, used to modify the calibration of
Calibration a data file, see the “To Modify a Calibration” section, on page 7-97.

Lock Mass Invokes the Lock Mass dialog; this allows the User to specify a mass that
will be located in the spectrum and used to calculate an offset that can be
applied to the rest of the spectrum. See the “Lock Mass” section, on
page 7-97 for details.

The Spectrum Window Menu

Figure 7.13 The Spectrum Window Menu

Tile Displays the current windows in a tiled view.

Cascade Displays the current windows in a cascaded view.

Stack Displays the current windows in a stacked view.

Arrange Icons Arranges the icons of minimized windows at the bottom of the
Chromatogram Window.

New Trace Invokes the New Spectrum dialog see the “The New Spectrum Dialog”
section, on page 7-22 for details.

Page 7-16
 Spectrum

List of current Click on the required trace to select it.

traces

The Spectrum Help Menu

The Help, Spectrum command invokes the Help function for Spectrum.

The Spectrum Tool Bar

General

The Spectrum Tool Bar is displayed at the top of the Spectrum Window. The default Spectrum
Tool Bar contains the buttons listed below. It is also possible to customize the Tool Bar and add
additional buttons for other Spectrum operations.

Tool Menu equivalent Purpose

Bar
button
File, Open Opens a data file.

File, Print Prints the current window in portrait format.

File, Print Prints the current window in landscape format.

Edit, Copy Picture Copies the current window to the clipboard.

Edit, Copy Spectrum Copies the currently-displayed range of the spectrum

List trace to the clipboard as mass and intensity pairs in the
form of a text list.

Edit, Paste Pastes the contents of the clipboard into the display.

Process, Refine Refines the current scan. The refine process identifies
the masses that contribute to a particular peak in the
TIC.

Tools, Library Search Identifies the current scan using the library search
facility.

Process, Process All Select once to process all traces in the current window.
Traces Select again to process only the current trace in the
current window.

Invokes the Edit Text String dialog; this allows text to

be added to a spectrum.

Selecting once causes each subsequent spectrum to

appear in a new window, rather than being added to the
current one. Selecting a second time cancels this mode.

Selecting once causes each subsequent spectrum to

replace the currently selected trace. Selecting a second
time cancels this mode.

Page 7-17
Spectrum

Note:

The button is grayed when the button is depressed.

Display, Real-Time Toggles real time spectrum data update on and off.
Update
Display, Range, Increases the magnification of the current range.
Magnify
Display, Range, Decreases the magnification of the current range.
Magnify
Display, Range, Deletes the current magnification range.
Magnify
Display, Spectrum, At Selects a new scan from the current data file.

Decrements the currently displayed scan.

Increments the currently displayed scan.

Select once to restore the previous display range; select

again to use the default display range.

Customizing the Spectrum Tool Bar

General

The Spectrum Tool Bar can be customized to:

• Add buttons for frequently used operations.

• Remove buttons that are not required.

• Change the order in which the Tool Bar buttons are displayed.

The additional buttons that can be added to the default Spectrum Tool Bar are:

Tool Menu equivalent Purpose

Bar
button
File, Save Spectrum Saves the spectrum.

Process, Smooth Invokes the Spectrum Smooth dialog.

Process, Combine Invokes the Combine Spectrum dialog.

Process, Subtract Invokes the Background Subtract dialog.

Process, Center Invokes the Spectrum Center dialog.

Process, Mass Measure Invokes the Mass Measure dialog.

Process, Component, Invokes the Manual Find Components dialog.

Find Manual

Page 7-18
 Spectrum

Tool Menu equivalent Purpose

Bar
button
Process, Component, Invokes the Automatic Find Components dialog.
Find Auto

Process, Transform Invokes the Transform dialog.

Tools, ACD Labs Invokes the Advanced Chemistry Development's

SpecManager ACD/Spec Manager software suite for structural
elucidation, (if installed on the PC).

Window, Tile Tiles the windows.

Window, Cascade Cascades the windows.

Window, Stack Stacks the windows.

The Customize Toolbar dialog

To customize the Spectrum Tool Bar, select the Spectrum Menu Bar Display, Customize Toolbar
command; the Customize Toolbar dialog is invoked.

Figure 7.14 The Customize Toolbar dialog

Available This list box contains all the available buttons that are not currently in the
Buttons: list box Tool Bar. A button can be selected by clicking on it.

The top entry in the box is Separator; it is never removed from the
Available Buttons: list box. However, it can be added to the Toolbar
Buttons list box to insert a separation gap between the buttons in the Tool
Bar.

Tool Bar This list box contains all the buttons that are currently in the toolbar. A
Buttons: list box button can be selected by clicking on it. The last entry in this box is
always Separator (dimmed); it cannot be removed from the list box, it
allows buttons to be added to the end of the list.

Add-> Moves the selected button from the Available Buttons: list box to the
Tool Bar Buttons: list box.

Page 7-19
Spectrum

<-Remove Moves the selected button from the Tool Bar Buttons: list box to the
Available Buttons: list box.

Note:
This button is grayed if no item is selected in the Tool Bar Buttons: list
box.

Close Exits the Customize Toolbar dialog.

Reset Resets the Tool Bar to its default display.

Move Up Moves the selected button one position up the list in the Toolbar Buttons:
list box.

Note:
This button is grayed if no item is selected in the Tool Bar Buttons: list
box, or if the top item in the list is selected.

Move Down Moves the selected button one position down the list in the Toolbar
Buttons: list box.

Note:
This button is grayed if no item is selected in the Tool Bar Buttons: list
box, or if the bottom item in the list is selected.

To Add Buttons to the Tool Bar

1. Select the Spectrum Menu Bar Display, Customize Toolbar command; the Customize
Toolbar dialog is invoked.

2. Select the button to be added in the Available Buttons: list box.

3. Select the Tool Bar button before which the new button is to be added in the Toolbar
Buttons: list box.

4. Select the Add button. The new button is added to the Toolbar Buttons: list box.

5. Repeat steps 2 to 4 to add further buttons to the Tool Bar.

6. Separators can be inserted between Tool Bar buttons to divide them into logical groups. To
add a separator, repeat steps 2 to 4 selecting Separator in the Available Buttons: list box.

7. Select the Close button to exit and save the changes.

To Remove Buttons from the Tool Bar

1. Select the Spectrum Menu Bar Display, Customize Toolbar command; the Customize
Toolbar dialog is invoked.

2. Select the button to be removed in the Toolbar Buttons: list box.

3. Select the Remove button. The button is removed from the Toolbar Buttons: list box.

4. Repeat steps 2 and 3 to remove further buttons from the Tool Bar.

5. Select the Close button to exit and save the changes.

Page 7-20
 Spectrum

To Change the Order in which Tool Bar Buttons are Displayed

1. Select the Spectrum Menu Bar Display, Customize Toolbar command; the Customize
Toolbar dialog is invoked.

2. Select the button to be moved in the Toolbar Buttons: list box.

3. Select the Move Up or Move Down buttons to move the Tool Bar button.

4. Repeat steps 2 and 3 as often as required.

5. Select the Close button to exit and save the changes.

To Reset the Tool Bar to the Default Settings

1. Select the Spectrum Menu Bar Display, Customize Toolbar command; the Customize
Toolbar dialog is invoked.

2. Select the Reset button.

3. Select the Close button to exit and save the changes.

To Remove the Tool Bar from the Spectrum Display

Select the Menu Bar Display, Toolbar command, the Tool Bar will be removed from the display.
A tick mark appears next to this menu item when it has been selected.

To re-display the Tool Bar, select the Menu Bar Display, Toolbar command again.

Displaying Spectra
Adding or Replacing Spectra

MassLynx provides a number of options for displaying new spectrum traces. New spectrum traces
can be generated by

• Opening a new file.

• Processing spectra (subtract, smooth, center, etc.).

• Selecting spectra by double-clicking on a chromatogram.

To display each new spectrum trace in a new window, select the Tool Bar button. To cancel
this mode and display new traces in the existing window select the Tool Bar button again.

When a new trace is displayed in the existing window, it can be added to the traces currently
displayed, or it can replace the current trace. Select the Tool Bar button once to cause each
subsequent spectrum, or spectrum process, to replace the currently selected trace. Selecting the
button a second time causes each subsequent spectrum, or spectrum process, to be added to the
traces on display. Up to sixteen spectrum traces can be displayed in one window.

Note:

1. The button is grayed when the button is depressed.

Page 7-21
Spectrum

2. The manner in which spectra are added to the Spectrum Window can also be selected via the
Menu Bar Window, New Trace command, refer to the “The New Spectrum Dialog” section,
below, for details.

The New Spectrum Dialog

The New Spectrum dialog is used to select the manner in which spectra are added to the Spectrum
Window; it is invoked by the Menu Bar Window, New Trace command.

Figure 7.15 The New Spectrum dialog

Add Trace Adds the spectrum to the current Spectrum Window.

Replace Trace The spectrum replaces the currently selected spectrum in the Spectrum
Window.

New Window Displays the spectrum in a new Window.

Viewing a Peak List Entry

To view a Peak List entry, select the Menu Bar Display, Spectrum, Peak List Entry command,
this invokes the Display Quan DB Spectrum dialog.

Figure 7.16 The Display Quan DB Spectrum dialog

File: Displays the current file.

Entry: Enter the required entry number, this field will only accept integer values
in the range 1 to the number of entries in the Peak List.

File Invokes the standard windows file Open dialog.

Add Trace Adds the spectrum to the current Spectrum Window.

Replace Trace The spectrum replaces the currently selected spectrum in the Spectrum
Window.

New Window Displays the spectrum in a new Window.

Page 7-22
 Spectrum

Manipulating the Display

Altering the Range of the Mass Axis

To Alter the Range of the Mass Axis using the Mouse

Click and hold the left mouse button at one end of the region of interest and drag the cursor
horizontally to the other end. As the cursor is dragged, a “rubber band” is stretched out to indicate
the range selected; do not go beyond the bounds of the axis. When the mouse button is released,
the selected range will be re-displayed to fill the current window.

This operation can be repeated as often as required.

To Alter the Range of the Mass Axis using the Menu Bar

1. Select the Menu Bar Display, Range, From command. The Display Range dialog is
invoked.

Figure 7.17 The Display Range dialog

2. Enter new From and To values for the mass axis.

3. Select the OK button.

Altering the Range of the Intensity Axis

Click and hold the left mouse button at one end of the region of interest and drag the cursor
vertically to the other end. As the cursor is dragged, a “rubber band” is stretched out to indicate
the range selected; do not go beyond the bounds of the axis. When the mouse button is released,
the selected range will be re-displayed to fill the current window.

This operation can be repeated as often as required.

Altering the Range of Both Axes

Click and hold the left mouse button at one end of the region of interest and drag the cursor to the
diagonally opposite corner. As the cursor is dragged, a “rubber band” is stretched out to indicate
the region selected; do not go beyond the bounds of the axes. When the mouse button is released,
the selected region will be re-displayed to fill the current window.

This operation can be repeated as often as required.

Setting Magnified Ranges

Setting Magnification Ranges using the Mouse

Click and hold the middle mouse button at one end of the region of interest and drag the cursor
horizontally to the other end. As the cursor is dragged, a “rubber band” is stretched out to indicate

Page 7-23
Spectrum

the range selected; do not go beyond the bounds of the axis. When the mouse button is released,
the selected range will be re-displayed with an initial magnification factor of two.

Alternatively, pressing the Shift key while using the left mouse button will perform the same
operation.

Setting Magnification Ranges using the Menu Bar Magnify Command

1. Either:

a. Select the Menu Bar Display, Range, Magnify command.

Or:

b. Double-click on the range magnification description of an existing magnified range.

In either case, the Spectrum Magnify dialog is invoked.

Figure 7.18 The Spectrum Magnify dialog

2. Enter the magnification factor to be applied in the By text box.

3. Enter the range to be magnified in the From and To text boxes.

4. To define more than one magnification range on the displayed Spectrum, select a new range in
the Range list box and repeat Steps 2 and 3. Up to five different magnified regions of the
Spectrum can be defined.

5. Select the OK button to close the dialog. The Spectrum is re-displayed with the data in the
selected regions magnified by the requested factor. The magnified regions are displayed in a
different color and labeled with the magnification factor.

Where multiple magnification regions have been defined, to select the current magnification range,
click in the magnification description that appears above the range. The description will change
color to red to indicate the currently selected range.

To Magnify the Range of the Intensity Axis using the Tool Bar

Select to increase the magnification of the current range. The current magnification
factor is multiplied by 1.5, and rounded up to the nearest even number to give the
increased magnification factor. If the initial magnification factor is 2, this will give
subsequent magnification factors of 4, 6, 10, 16, etc.

Select to decrease the magnification of the current range. The current magnification
factor is divided by 1.5, and rounded down to the nearest even number to give the
decreased magnification factor. If the initial magnification factor is 16, this will give
subsequent magnification factors of 10, 6, 4, etc.

Page 7-24
 Spectrum

To Change the Magnification of a Particular Range

1. Either:

a. Select the Menu Bar Display, Range, Magnify command.

Or:

b. Double-click on the range magnification description of an existing magnified range.

In either case, the Spectrum Magnify dialog is invoked, see the “Setting Magnification
Ranges using the Menu Bar Magnify Command” section, on page 7-24.

2. Enter the new magnification factor in the By text box.

3. Select the OK button.

Deleting Magnification Ranges

Select the Tool Bar button to delete the current magnification range.

To delete all the magnification ranges:

1. Either:

a. Select the Menu Bar Display, Range, Magnify command.

Or:

b. Double-click on the range magnification description of an existing magnified range.

In either case, the Spectrum Magnify dialog is invoked, see the “Setting Magnification
Ranges using the Menu Bar Magnify Command” section, on page 7-24.

2. Select the Default button; this will delete all magnification ranges.

3. Select the OK button.

Restoring the Display

Selecting the Tool Bar button once restores the display to its previous state. Selecting it a
second time restores the display to the default range.

Note:
These operations do not remove magnification ranges.

Page 7-25
Spectrum

Setting the Display Range Defaults

Note:

The display range default settings specify both the effects of selecting the Tool Bar button, and
adding a new Spectrum to the display.

To Change the Default Display Range

1. Select the Menu Bar Display, Range, Default command; the Default Spectrum Range
dialog is invoked.

2. Make the required changes, see below.

3. Select the OK button.

Figure 7.19 The Default Spectrum Range dialog

Default range Specifies whether the mass axis will range from the first peak to the last
Frame peak in the scan (Data), or over the range requested when the acquisition
started (Acquisition).

Note:
This frame is only relevant to Centroid mode acquisitions.

Default graph If there is more than one spectrum in a window, this option specifies
Frame whether the mass range for that window is made large enough to include
the mass ranges of All the spectra, or large enough for the Current
spectrum only.

Automatic range If this option is checked, the display range will return to the specified
default default (see Default range and Default graph above) when a new
spectrum is added to a Spectrum Window. If this option is not checked,
the display range will remain unchanged when a new spectrum is added.

Displaying a Spectrum as a List

General

The display in the current spectrum window can be replaced with a list of masses and intensities of
the peaks in the currently selected spectrum.

To Display a Spectrum as a List

Select the Menu Bar Display, List Spectrum command. A check mark is placed against the List
Spectrum menu item. Most of the Menu Bar commands and the Tool Bar may still be used.

Page 7-26
 Spectrum

Figure 7.20 Typical spectrum displayed as a list

To Restore the Graphical Display

Select the Menu Bar Display, List Spectrum command. The check mark is removed from the
List Spectrum menu item.

To Print a Report of the Spectrum Listing

1. Select the Menu Bar File, Print Report command. The Spectrum Print Report dialog is
invoked.

Figure 7.21 The Spectrum Print Report dialog

Page 7-27
Spectrum

2. Select the Range of data to be displayed. Select Data to print a listing of the whole data file.
Select Display to print a listing of the current display range.

3. Select the Header Information and Peak Information to be printed by selecting the relevant
check boxes.

4. Select the OK button to exit and print the report.

Controlling the Appearance of the Display

General

Each Spectrum Window has its own set of Display Parameters, which determine the appearance of
the Spectrum display. The parameters can be inspected and altered for the current Spectrum
Window from the Spectrum Display dialog.

To Change the Display Parameters

1. Select the Menu Bar Display, View command; the Spectrum Display dialog is invoked.

2. Make the required changes, see below.

3. Select the OK button.

Figure 7.22 The Spectrum Display dialog

Page 7-28
 Spectrum

Normalize Data These controls specify the scale on the intensity axis.
To Frame
Largest Peak Displays the largest peak currently on display at 100% of the intensity
on Display axis.

Base Peak in Displays the largest peak at 100% of the intensity axis.
Spectrum
Mass When selected, 100% on the intensity axis represents the height of the
peak at the mass specified in the adjacent text box.

Intensity When selected, 100% on the intensity axis represents the intensity
specified in the adjacent text box.

Baseline at Scales the vertical axis from 0%.

Zero
Baseline When selected, the vertical axis is scaled from the intensity specified in the
adjacent text box. This option can be useful for displaying spectra that
have a raised baseline.

Link Vertical Gives all axes in the current window a common vertical scale. This
Axes enables two spectra to be plotted on the same intensity scale, in order to
overlay and compare them.

Data Threshold When processing centroid type data, it can be useful to specify an intensity
Frame threshold. Peaks whose intensity is less than the threshold will not be
displayed. This frame allows this to be done.

Note:
The threshold controls are not applicable to continuum mode data.

% Full scale Sets the threshold as a percentage of the intensity of the largest peak in the
spectrum.

Intensity Sets an absolute intensity threshold.

Style Frame

Overlay Allows multiple traces in the same window to be superimposed on the

Graphs same axis.

If the option is not selected, the traces will be drawn on separate axes,
arranged vertically.

Note:
When spectra are overlaid, only the currently selected trace is annotated.

Graph Header Displays the graph header information at the top of the Spectrum.

Page 7-29
Spectrum

Process Displays process information in the Spectrum header.

Description
Note:
The Graph Header option overrides the Process Description option, i.e.
if the Graph Header is deselected, the Process Description will also be
deselected.

Component Displays a summary of the components identified so far on each spectrum.

Table
Show Negative The Background Subtract process sets a zero level in continuum data, and
Data in the resultant spectrum, half the noise lies below that level. This option
specifies whether these negative data points are displayed. If the option is
selected, the scale on the intensity axis ranges from the smallest (most
negative) intensity to the largest. If the option is not selected, the intensity
axis ranges from zero to the largest intensity. Refer to the “The
Background Subtract Process” section, on page 7-40, for further details.

Show Zero Draws a horizontal line to represent the zero level in the spectrum. Again,
Level this is useful for gauging the effect of the Background Subtract process.

Hide Lock For Mass Measured Tof data a Lock Mass Peak can be defined, this peak
Mass Peaks will be shown in a different color on the spectrum. This option specifies
whether the lock mass peak is displayed. If the option is selected, the
Lock Mass Peak is not displayed. Refer to the “Lock Mass” section, on
page 7-97, for further details.

Fill Trace Colors the area under the spectrum.

Note:
This option only applies to continuum-type (not centroid) data.

Split Axis This option is enabled when the Overlay Graphs control is selected. It
allows the User to alter the aspect ratio of the spectrum by dividing the
mass axis into segments, then arranging the segments vertically. For
example, if a spectrum of from 40 to 340 amu is on display, and 3 is
selected in the Split Axis option, the display will show three axes: one
from 40 to 140 amu, one from 140 to 240 amu, and one from 240 to
340 amu.

Overlay This option is enabled when the Overlay Graphs control is selected. It
Step X(%) allows the User to offset each subsequent spectrum trace by a percentage
of the horizontal axis. This can make it easier to examine overlaid traces.

Overlay This option is enabled when the Overlay Graphs control is selected. It
Step Y(%) allows the User to offset each subsequent spectrum trace by a percentage
of the intensity axis. This can make it easier to examine overlaid traces.

Page 7-30
 Spectrum

Grid Enables the User to specify a grid to be displayed on the Spectrum display.
The pattern of the lines that make up the grid can be chosen as Dot, Dash,
or Solid. Select Off if no grid is to be displayed.

Header button Invokes the Header Editor, which allows editing of the header
information displayed at the top of the window. For more information, see
the “The Header Editor Dialog” section in Chapter 3, “The MassLynx
Window and Related Information”.

Controlling the Appearance of Peak Labels

General

Each Spectrum Window has its own set of Peak Annotation Parameters, which determine the
appearance of peak labels. The User can inspect and alter the parameters for the current window
in the Spectrum Peak Annotation dialog.

To Change the Peak Annotation Parameters

1. Select the Menu Bar Display, Peak Annotation command; the Spectrum Peak Annotation
dialog is invoked.

2. Make the required changes, see below.

3. Select the OK button.

Figure 7.23 The Spectrum Peak Annotation dialog

Page 7-31
Spectrum

Annotation Type
Frame
Decimal Places Select between zero and four decimal places to be displayed on mass
labels.

Note:
This control does not affect intensity labels, which are always displayed as
integers.

Mass Labels peaks in the current spectrum window with their masses to the
specified number of decimal places.

Intensity Labels peaks in the current spectrum window with their intensity as an
integer value.

Delta Mass Displays the difference between the mass of each peak in the spectrum and
the specified mass.

Note:
The following controls are only applicable to ElectroSpray data.

Mass Error Labels peaks in a MaxEnt Spectrum with their probable mass errors.

Intensity Error Provides an error range on the intensity of MaxEnt Spectrum peaks.

Component Labels peaks with the name of the appropriate component, and charge state
Label if raw data is being viewed. (In Transform or MaxEnt spectra, peaks are
labeled with component name alone.)

Ion Series Enables the Series button; this invokes the Ion Series Annotation dialog,
Label which allows the User to select which ion series to annotate (see the “Mass
Spectral Fragments” section in the BioLynx & ProteinLynx User’s Guide).

Files with extensions ion, int and tab are created and stored in the raw data
file when matching theoretical mass spectral fragment ions generated in
BioLynx with centroided data.

Digest Annotates the digest fragments matched from BioLynx, e.g. T10-11.
fragment label

Annotation Used to enter the minimum intensity for a peak to be labeled.

Threshold frame
% Full scale When selected, enter a value in the adjacent text box to define the
threshold as a percentage of the base peak intensity.

Intensity When selected, enter a value in the adjacent text box to define the
threshold intensity.

Level From the drop down list box, select the number of labels that appear on the
chromatogram; this can be set to High, Medium or Low.

Page 7-32
 Spectrum

To Annotate a Particular Peak

Hold down the keyboard Ctrl key and right-click on the peak to be annotated. The peak will be
mass labeled.

To remove the mass label from the peak, hold down the Ctrl key and right-click on the peak a
second time.

Removing Spectra from the Display

To Remove a Single Spectrum Trace from the Display

1. Press the keyboard Delete key. A dialog is invoked asking for confirmation of deletion of the
currently selected spectrum trace.

Figure 7.24 The Short Cut Remove Spectrum dialog

2. Select the OK button; the dialog is closed and the selected traces are removed from the
display. This operation does not affect the data stored on disk.

To Remove Multiple Spectrum Traces from the Display

1. Select the Spectrum Menu Bar Display, Remove command. The Remove Spectra dialog is
invoked.

Figure 7.25 The Remove Spectra dialog

2. The spectra in the current window are listed in the order in which they appear on the display.
One or more spectra can be selected by clicking in the Spectra: list box. Clicking again on a
selected item will cancel the selection. Selecting the All button selects all the spectra.

3. Select the OK button; the dialog is closed and the selected trace(s) are removed from the
display. This operation does not affect the data stored on disk.

Real-Time Display of Spectra

If data are being acquired into a file, the associated spectra can be displayed in real time, by
selecting the Tool Bar button; or the Spectrum Menu Bar Display, Real-Time Update
command, which invokes the Spectrum Real-Time Update dialog.

Page 7-33
Spectrum

Figure 7.26 The Spectrum Real-Time Update dialog

Enable Real- Enables the Real-Time update.

Time update
Update Frame

Latest scan When selected, displays the latest scan.

Average all When selected, displays the average of all the scans acquired at present.
scans
Average When selected, displays the average of the latest number of scans defined
latest…scans in the text box.

Each spectrum window has a separate real time update switch. The state of the switch for a
particular window can be ascertained by checking if the Tool Bar button is depressed, or by
checking the state of the Spectrum Real-Time Update dialog Enable Real-Time update option.

Changing the Order of Displayed Spectra

When a window contains multiple traces, the order in which they are displayed can be changed.
The spectrum that is first in the list is displayed at the bottom of the screen.

Select the Spectrum Menu Bar Display, Move To First option to display the currently selected
spectrum at the bottom of the screen.

Select the Spectrum Menu Bar Display, Move To Last option to display the currently selected
spectrum at the top of the screen.

Adding Text to the Spectrum Display

User text labels are added to a spectrum display in an identical manner to that for Chromatogram,
refer to the “Adding Text to the Chromatogram Display” section, in Chapter 6, “Chromatogram”
for details.

Exporting SEQUEST Files

General

MassLynx has a facility to convert files into a format that can be used by the “SEQUEST”
program. The “SEQUEST” program correlates uninterpreted tandem mass spectra of peptides

Page 7-34
 Spectrum

with amino acid sequences from protein and nucleotide databases. It has been written by Jimmy
Eng and John Yates (University of Washington).

Note:
This option is only enabled if BioLynx is installed.

To Export a SEQUEST File

1. Display the relevant centered MS/MS data file.

2. Select the Menu Bar File, Export SEQUEST file command. The Export SEQUEST
compatible file dialog is invoked.

3. Make the required changes, see below.

4. Select the OK button.

Figure 7.27 The Export SEQUEST compatible file dialog

Precursor ion The precursor ion mass is picked up from the data file, if it was entered in
mass: the Function Editor, otherwise, a value may be entered in this text box.

Precursor charge This value defaults to 2, it may be changed as required.

state:
Partial sequence Any known sequence information may be entered in this box.
information:
File Path and The location and file name that the file will be saved to. The file name is
Destination: the original file name, with the scan and function numbers appended to it.
To change the destination, type a new destination in this box, or select the
Browse button and select a new destination from the displayed dialog.

Processing Spectra
General

Several processes are available for use on spectra:

• Refine, see the “The Refine Process” section, on page 7-38.

• Combine Spectra, see the “The Combine Spectra Process” section, on page 7-39.

Page 7-35
Spectrum

• Background Subtract, see the “The Background Subtract Process” section, on page 7-40.

• Smooth, see the “The Smooth Process” section, on page 7-41.

• Center, see the “The Center Process” section, on page 7-43.

• Mass Measure, see the “The Mass Measure Process” section, on page 7-46.

• Integrate, see the “The Integration Process” section, on page 7-48.

Saving and Recalling Processed Spectra

General

The spectra resulting from any spectral processing can be saved with the raw data.

To Save a Processed Spectrum

Select the processed spectrum in the Spectrum Window and select the Menu Bar File, Save
Spectrum command.

The Spectrum Save dialog is invoked, giving a brief description of the process being saved.
Select the OK button to save the process and close the dialog.

Figure 7.28 The Spectrum Save dialog

To Reload Processed Data into Spectrum

1. Select the Menu Bar File, Open command; the Spectrum Data Browser dialog is invoked.

Note:
The Spectrum Data Browser dialog is based on the standard MassLynx Data Browser dialog,
see the “Opening Data Files: The MassLynx Window Data Browser Dialog” section, in
Chapter 3, “The MassLynx Window and Related Information” for details.

Page 7-36
 Spectrum

Figure 7.29 The Spectrum Data Browser dialog

2. Click on the raw data file from which the processed data was obtained.

3. Select the History button; the History Selector dialog is invoked, see the “The History
Selector Dialog” section, in Chapter 3, “The MassLynx Window and Related Information” for
details.

Figure 7.30 The History Selector dialog

Page 7-37
Spectrum

4. In the Process History: list, select the processed data to be loaded.

5. Select the OK button to exit the History Selector dialog.

6. Select the OK button to exit the Spectrum Data Browser dialog and load the processed data.

The Refine Process

General

The Refine process operates on centroid-mode data only. Its purpose is to automatically remove
background ions from a spectrum thereby allowing it to be more easily identified, for example by
library search.

Select the Menu Bar Process, Refine command to invoke the Refine Spectrum dialog.

Figure 7.31 The Refine Spectrum dialog

A particular TIC peak is identified by specifying the peak-top scan. The User supplies two
parameters for the process; Window size and Noise threshold.

The Refine algorithm proceeds by generating the summed mass chromatogram over a range of
1 Da centered on each integer mass in turn. It examines these chromatograms for a number of
scans equal to the Window size around the peak top scan. (Window size is the half width in
scans at baseline of the TIC peak of interest.) If there is a peak present in this range whose top-
most point is within one scan of the peak top scan, and is more intense than the Noise threshold
value, then this mass will appear in the refined spectrum.

To Refine a Scan in a Centroid-Mode Data File

1. Identify the scan at the top of the peak of interest. Display this scan in a spectrum window.
This can be simply done by double-clicking on the peak in the Chromatogram Window.

2. Select the Spectrum Menu Bar Process, Refine command, the Refine Spectrum dialog is
invoked.

3. Enter values for Window size and Noise threshold. For the first run, set Noise threshold to
zero to show all peaks.

4. Select the OK button to start the process.

5. If the noise level in the refined spectrum is unacceptable, repeat the refine operation with a
higher Noise threshold setting. Values in the range 0 to 10 are recommended.

The current spectrum may also be refined, using the current refine parameters, by selecting the
Spectrum Tool Bar button.

Page 7-38
 Spectrum

The Combine Spectra Process

General

The Combine Spectra process operates on centroid-mode or continuum data. Its purpose is to
produce a single spectrum by subtracting averaged background spectra from the average of spectra
produced by multiple scans of a single TIC peak. The combined spectrum exhibits enhanced
signal-to-noise and improved mass accuracy.

The User specifies three scan ranges and a background factor. One range contains the scans across
the peak top (the peak-top scan range) and the other two ranges contain scans from the
background, on each side of the peak. The scans across the peak top are averaged together and the
average of all the background scans, multiplied by the background factor (X), is subtracted from
the result.

To Combine Scans in a Centroid-Mode Data File

1. Display the chromatogram peak of interest in a Chromatogram Window.

2. Select the Tool Bar button, or select the Menu Bar Process, Combine spectra command;
the Combine Spectrum dialog is invoked.

Note:
The Combine Spectrum dialog may also be invoked from a Chromatogram Window by selecting
the Chromatogram Menu Bar Process, Combine Spectra command.

3. Select the desired options (see below).

4. Select the OK button, the Combine Spectra process starts.

Figure 7.32 The Combine Spectrum dialog

Average Specifies the peak-top scan range. This can be entered either by typing
scan numbers separated by a colon (e.g. 619:626) in the text box, or by
ensuring that the focus is in the text box and then dragging across the peak
using the right mouse button.

Note:
This field will only accept scan numbers in the range of the appropriate
raw data file.

Peak separation Specifies the maximum resolution of peaks in amu. This determines
which peaks are to be regarded as being due to the same peak from scan to
scan.

Page 7-39
Spectrum

Subtract Specifies the background scan range(s), that is, each side of the peak.
These can be entered by typing scan numbers in the text box; each range
must be in the form of two numbers separated by a colon. If there are two
ranges, they must be separated by a comma (e.g. 606:612,631:637).
Alternatively, the range can be entered by ensuring that the focus is in the
text box and then dragging across a sample of the background using the
right mouse button.

Note:
This field will only accept scan numbers in the range of the appropriate
raw data file.

Multiple Normally, when using the right mouse button to enter values, the first set
Average of ranges is entered in the Average box and the second and third sets are
entered in the Subtract box. Selecting the Multiple Average box changes
this so that the first six sets of ranges are entered in the Average box and
the seventh and eighth are entered in the Subtract box.

X Specifies the background factor; a value of 1 is equivalent to no scaling.

Values less than or equal to zero will default to 1.

Reset Clears the Average and Subtract text boxes.

The Background Subtract Process

General

Background Subtract adjusts the zero level in a continuum spectrum to lessen the effect of
chemical noise caused by column bleed, etc.

Both the Transform and MaxEnt processes rely on having background removed from the
spectrum; MaxEnt, especially, will produce an inferior result if this is not done. On data with a
curved background, typically ElectroSpray and FAB spectra, Background Subtract improves
presentation and aids interpretation.

A low order polynomial is fitted to the data to remove a constant, sloping or curved background
from a spectrum. The algorithm fits a polynomial of specified order (zero is a flat baseline, one is
a straight, sloping line, two is a quadratic shape, etc.) to a spectrum, such that a specified
percentage (usually 30 to 50%) of the data points lies below the polynomial. This operation is
performed to an arithmetical tolerance that is specified by the User.

The Background Subtract process also gives the User the option to display a graph of the baseline,
which will be fitted to the data before starting the process.

To Subtract the Background from a Continuum Spectrum

1. Select the Menu Bar Process, Subtract command; the Background Subtract dialog is
invoked.

2. Select the desired options (see below).

3. Select the OK button, the Combine Spectra process starts. The Subtract status dialog box
indicates the progress of the subtract algorithm.

The convergence value in the dialog box is updated after every iteration. The algorithm
terminates when convergence is less than tolerance. The User can choose whether to view

Page 7-40
 Spectrum

the zero level and negative data in the spectrum by selecting the appropriate options in the
Spectrum Display View dialog.

Figure 7.33 The Background Subtract dialog

Polynomial Specifies the order for the polynomial: 0 is a flat baseline, 1 is a straight,
order sloping line, 2 is a quadratic shape, etc.

Below curve (%) Specifies the percentage of data points that lie below the polynomial. The
effect of increasing this parameter is to raise the zero level in the spectrum.
The default value of 40% is based on the observation that around 80% of
the data points in a typical ElectroSpray spectrum are noise, and only 20%
signal. Half the noise lies above the zero line, and half below, therefore
half of 80%, or 40% of the total number of data points, should lie below
the background zero level.

Tolerance The effect of increasing this parameter is to make the algorithm terminate
sooner, but the result may not be as satisfactory.

Flatten edges When selected, the software checks that the applied polynomial is flat or
horizontal at the beginning and end of the trace.

Make graph of Gives the User the option of seeing what the effect of the Background
fitted polynomial Subtraction would be on the data before actually doing it. Select this
option, then select the OK button. A graph of the polynomial function,
which would be subtracted from the spectrum, is displayed above the
resulting subtracted spectrum. If the Spectrum Display dialog Link
Vertical Axes and Overlay Graphs options are selected, the new baseline
will be superimposed on the existing data. When satisfied with the
parameters being used, deselect the Make graph of fitted polynomial
option.

The Smooth Process

General

Smoothing reduces the high-frequency noise present in a spectrum, thus aiding interpretation. It is
strongly recommended that data is smoothed before mass measurement is attempted with the
Center process, otherwise peaks may be created from the noise spikes.

Note:
Data for MaxEnt must not be smoothed.

Three types of smoothing are implemented in MassLynx for smoothing spectra:

• Moving Mean.

Page 7-41
Spectrum

• Savitzky Golay.

• Moving Median.

The most useful technique is Moving Mean. Using Savitzky Golay allows a heavier smooth
without broadening the peak as much. Moving Median is used for removing noise spikes that are
very much narrower than the real peaks (single ions, impulses from the electronics, etc.).

All three smoothing methods slide a window along the data, averaging the data in the window to
produce a point in the smoothed spectrum. The width of the smoothing window, in data points, is
determined by the data system using the equation:

Full peak width at 50% intensity

Halfwidth of smoothing window = ,
3δm

where δm is the spacing between adjacent points on the mass axis, i.e., 0.0625 Da for raw
continuum/MCA data, or equal to the value of the Resolution parameter for MaxEnt or Transform
data.

Moving Mean takes the arithmetical mean of the intensities of the data points in the window.

Savitzky Golay takes an average of the intensities weighted by a quadratic curve. This tends to
enhance quadratic-shaped features in the data (peaks).

Moving median takes the arithmetical median of the intensities of the data points in the window.
This process is unlike the previous two in that the median smooth iterates until the spectrum no
longer changes. The effect is that the intensity of narrow spikes is reduced on successive
iterations, to background level on convergence.

To Smooth a Continuum Spectrum

1. Expand a section of the spectrum sufficient to allow an estimate to be made of the width of a
peak at half height.

2. Choose the Menu Bar Process, Smooth command, the Spectrum Smooth dialog is invoked,
see below.

3. Set the Peak width (Da) parameter according to the value estimated in step 1, this can be
done by dragging, using the mouse, over the peak at half height.

4. If Moving Mean or Savitzky Golay have been selected, the number of times the smooth is
repeated may be changed, by changing the Number of smooths parameter from its default
value of 2. Increasing this parameter gives a heavier smooth.

5. Select the OK button, the Smooth process starts.

Figure 7.34 The Spectrum Smooth dialog

Page 7-42
 Spectrum

Peak width (Da) An estimate for this parameter may be obtained by dragging, using the
mouse, over the peak at half height.

Number of Specifies the number of times the smooth is repeated; increasing this
smooths parameter gives a heavier smooth. The maximum value is 100.

Note:
The Number of smooths parameter has no effect on Median smoothing,
which always iterates until the spectrum is unchanged.

Smoothing
method Frame
Mean Selects the Moving Mean smoothing method.

Median Selects the Median smoothing method.

Note:
The Median smoothing algorithm has the side effect of producing peaks
with flattened tops. For this reason, it is recommended that a Median
smooth be followed by a single iteration of a Mean or Savitzky Golay
smooth.

Savitzky Golay Selects the Savitzky Golay smoothing method.

The Center Process

General

Peak centering uses all the points across a peak in a continuum trace to calculate the mass of the
peak center. The centering process can be used either to label each peak with the calculated mass,
or to produce a single bar from each peak in a continuum spectrum. The calculation can be
performed in three ways:

• Select the most intense (top) point on the peak. This method is the least prone to errors
caused by unresolved adducts in ElectroSpray spectra.

• Calculate the centroid of the peak. This is equivalent to finding the vertical line passing
through the center of gravity of the peak. This will provide a more accurate mass
measurement, unless the peak contains unresolved adducts.

• Calculate the median of peak area. This is equivalent to drawing the vertical line such that
half the area of the peak lies on either side.

There is little practical difference between the median and centroid methods, though it may be the
case that the median is a more robust statistic on very asymmetric peak shapes. Masses from
different experiments obtained by centering with different methods should not be compared.

The centering algorithm looks for the trace rising then falling to indicate the top of a peak. The
User specifies how many data points must be visible as a clear peak top before the algorithm turns
the peak into a bar.

Page 7-43
Spectrum

For the centroid method, there is also the option of only using a specified fraction of the resolved
part of the peak. This helps to avoid the mass given to the bar being affected by unresolved
neighboring peaks.

To Center a Continuum Spectrum

1. Background Subtract the spectrum, see the “The Background Subtract Process” section, on
page 7-40. Background subtraction tells the centering algorithm how much of the spectrum is
noise, and therefore reduces the amount of noise seen in the resultant bar spectrum.

2. Smooth the spectrum, see the “The Smooth Process” section, on page 7-41. Smoothing helps
the centering algorithm make sensible decisions about whether groups of data points represent
peaks, or noise spikes.

Note:
MaxEnt spectra are an exception; they need centering to get an accurate mass, just like any
continuum spectrum. However, MaxEnt is designed to produce smooth spectra, and every peak in
the MaxEnt result has already been interpreted by MaxEnt as significant. Hence, neither
smoothing nor subtraction of MaxEnt spectra is necessary prior to mass measurement.

3. Select the Menu Bar Process, Center command, the Spectrum Center dialog is invoked.

4. Select the desired options (see below).

5. Select the OK button, the Center process starts.

Figure 7.35 The Spectrum Center dialog

Page 7-44
 Spectrum

Center method
Frame
Min peak An estimate for this parameter may be obtained by dragging, using the
width at half mouse, over the peak at half height; alternatively, enter a value in the text
height box.
(channels)
This parameter determines how many data points must be visible in the
expected shape across the peak top, i.e. minimum width. For
continuum/MCA data, setting this parameter to 4 is safe. Since there are
sixteen data points collected per Dalton, the value 4 is equivalent to
0.25 Da. For MaxEnt results, the peaks can be very narrow. Sometimes
there are two data points across the peak top. Therefore, for MaxEnt
results, the only safe value for this parameter is 2.

Too low a setting of this parameter will result in the centering algorithm
producing bars from the high-frequency noise.

Too high a setting of this parameter will result in the centering algorithm
misinterpreting many peaks to produce a single bar.

Top Selects the top method of processing.

Centroid top Selects the centroid method of processing. The fraction of the resolved
(%) portion of the peak that is used to calculate the centroid may be changed,
from its default value of 80%, in the adjacent text box; recommended
values are 60% to 95%.

Median Selects the median method of processing.

Centered
spectrum Frame
Create Creates a bar spectrum; the masses of the bars are calculated according to
centered the selected center method.
spectrum
Heights When selected, the height of a bar represents the intensity of the
continuum trace at the mass of the bar.

Areas When selected, the height of a bar represents the sum of the intensities of
the points across the peak in the continuum trace.

Add Adds the bar spectrum to the current Spectrum Window.

New window Displays the bar spectrum in a new Window.

Replace The bar spectrum replaces the currently selected spectrum in the Window.

Note:
For Tof data this dialog will have an extra button. Select this button to display the QTOF
Accurate Mass parameters dialog. For details, see the “QTOF Accurate Mass” section, on
page 7-47.

Page 7-45
Spectrum

The Mass Measure Process

General

The Mass measure process performs a combination of background subtraction, smoothing and
centering all in one command. Select the Spectrum Menu Bar Process, Mass Measure command
to invoke the Mass measure dialog.

Figure 7.36 The Mass measure dialog

Background subtraction takes place if the Background subtract control is checked. The Mass
Measure dialog gives access to the Polynomial order and Below curve (%) parameters which
are described in the “The Background Subtract Process” section, on page 7-40.

Mean Smoothing takes place if the Mean smooth control is checked. The Mass Measure dialog
gives access to the Peak width, Number of smooths, Mean and Savitzky Golay parameters
which are described in the “The Smooth Process” section, on page 7-41.

Peak Centering always takes place when the Mass measure process is used. The Mass Measure
dialog gives access to the Min peak width at half height, Top and Centroid top parameters
which are described in the “The Center Process” section, on page 7-43.

The Mass measure dialog always retains the last set of parameters used.

Note:
For Q-Tof data the Mass measure dialog will have an extra TOF button; this invokes the TOF
Accurate Mass dialog. For details, see the “QTOF Accurate Mass” section, on page 7-47.

Page 7-46
 Spectrum

QTOF Accurate Mass

For Q-Tof data the Mass Measure dialog has a Use QTOF mass correction button which
invokes the TOF Accurate Mass dialog.

Figure 7.37 The TOF Accurate Mass dialog

TOF Constants
Frame
Resolution Enter the resolution of the Mass Spectrometer.

Np multiplier Enter a value for the Number of Pushes correction factor.

Lock Mass
Correction
Frame
Mass Window Determines the width of the mass window used to locate the lock mass
+/- data peak. The most intense peak in the range Lock Mass – Mass
Window to Lock Mass + Mass Window is selected, and mass correction
based on this peak is performed.

Lock Mass Specifies the reference lock mass, refer to the “Lock Mass” section, on
page 7-97, for further details.

The TOF Transform Process

Note:
The TOF Transform process is only available if the BioLynx or ProteinLynx application manager
has been installed.

The TOF Transform process works on centroided (normally Q-Tof data). It both de-isotopes
masses, and realigns to a single charge state mass axis. The TOF Transform dialog is invoked by
the Spectrum Menu Bar Process, TOF Transform command.

Page 7-47
Spectrum

Figure 7.38 The TOF Transform dialog

The values entered in the Min Molecular Mass and Max Molecular Mass text boxes specify the
mass range over which the final output data will be aligned. For example, if the original spectrum
has the largest mass of interest at 800 and the Max Charge State is 3, the mass range must be at
least 2400 (3 x 800). The Max Charge State value should not exceed 4. In this example, if a
mass at 700, with charge state 4, is present, it will not be seen (since 4 x 700 = 2800, and the
specified mass range is 2400).

The Integration Process

General

The Spectrum integration process locates spectral peaks, draws baselines and calculates peak
areas. Spectrum integration works over the full mass range of the spectrum.

The assignment of baselines and separation of partially resolved peaks by verticals is determined
by the Peak Detection parameters. For a detailed explanation of how the Peak Detection
parameters affect integration see the “Integrating Chromatograms” Section in Chapter 6,
“Chromatogram”.

To annotate the integrated spectrum with peak areas, select the Spectrum Peak Annotation dialog
Intensity option, see the “To Change the Peak Annotation Parameters” section, on page 7-31.

To Integrate a Spectrum

1. Select the Menu Bar Process, Integrate command. The Peak Detect dialog is invoked, refer
to the “Standard Peak Detection Parameters” section, in Chapter 6, “Chromatogram” for
further information.

2. Edit the Peak Detection parameters as required.

3. Select the OK button to exit the dialog and perform the integration. The integration software
will locate the peaks, draw baselines and calculate peak areas.

Page 7-48
 Spectrum

Figure 7.39 The Peak Detect dialog

ElectroSpray Data Processing

General

In the ElectroSpray spectra of proteins, etc, each component produces a range of multiply charged
ions in the original m/z spectrum. Therefore, additional processing must be performed to produce
a molecular mass spectrum, also, due to the high accuracy required, a special calibration procedure
is used.

MassLynx provides two distinct methods for calculating the molecular mass spectrum:

• Transform

In this process, the User assigns charge states to peaks in the ElectroSpray m/z spectrum. This
information is then used to transform the ElectroSpray data onto a molecular mass axis, see
the “The Transform Process” section, on page 7-58, for further information.

• MaxEnt

The MaxEnt algorithm uses the maximum entropy method to produce true molecular mass
spectra from multiply-charged ElectroSpray spectra, see the “MaxEnt 1” section, on
page 7-59, for further information.

Page 7-49
Spectrum

Setting Adduct Mass for Transform and MaxEnt

Both the Transform and MaxEnt processes use the value for adduct mass in their calculations. To
set the adduct mass value, select the Menu Bar Process, Set Adduct Mass command; this invokes
the Set Adduct Mass dialog. The Adduct Type can be set to Hydrogen, Potassium or Sodium.
Selection of more than one adduct type is not supported.

Figure 7.40 The Set Adduct Mass dialog

Finding Components for Transform

Initial Processing

Transform initially requires the assignment of charge states, and this is performed on a bar
spectrum. Therefore, the first three steps are:

1. Background subtract the data, refer to the “The Background Subtract Process” section, on
page 7-40. In the Background Subtract dialog (see Figure 7.33, on page 7-41), suggested
parameter values are: Polynomial order set to 1 for a flat baseline, or 5 for a curved baseline,
Below curve set to 40%, and Tolerance set to 0.010.

2. Smooth the data with the Moving Mean algorithm, refer to the “The Smooth Process” section,
on page 7-41. The width of a peak in the raw data at half its maximum intensity must be
measured; enter this value in the Spectrum Smooth dialog Peak width (Da) field, see
Figure 7.34, on page 7-42. Set the Number of smooths parameter to 2.

3. Create a bar spectrum with the Center process, refer to the “The Center Process” section, on
page 7-43. Set the Spectrum Center dialog Min peak width at half height (channels)
parameter to 4, see Figure 7.35, on page 7-44. Select Top as the centering method. Ensure
the Create centered spectrum and Heights options are selected. It is convenient to put the
bar spectrum into a new window, so it can be expanded to fill the Spectrum window when
multiply-charged series are being identified; select the New window option to do this.

Finding Components

Multiply-charged series can now be identified as components. There are two methods of
component identification:

• The manual method is used when knowledge about the expected component mass is
unknown. The User must identify two adjacent peaks in each series. MassLynx then
identifies the rest of the series above the threshold and calculates the component’s molecular
mass, and the standard deviation associated with this mass.

• The automatic method is used when knowledge about the expected component mass is
known. It can be used to find each series in the spectrum in turn, or to identify all series in the
spectrum. The disadvantage of this method is that a mass range to search over must be known
in advance. Using a wide mass range may result in the false identification of spurious series.

For the analysis of a true unknown, the manual method is preferred, so that the reliability of each
entry can be checked.

Page 7-50
 Spectrum

To Find Components using the Manual Method; the Manual Find Components Dialog

The manual method for finding components uses the Manual Find Components dialog.

Figure 7.41 The Manual Find Components dialog

Find Component
Frame
Peak 1 m/z and These parameters are entered by typing their values directly in the text
Peak 2 m/z boxes. Alternatively, after visually identifying a multiply charged peak
series, position the mouse pointer close to one peak in the series and right-
click. Position the pointer close to an adjacent peak in the series and right-
click again. The Peak 1 and Peak 2 controls will be updated to show the
selected masses.
Window… Da Specifies the tolerance on the position of each peak in the series. It may
need to be increased from its default value of 0.5 Da for statistically poor
data. Too low a value will result in the algorithm being unable to identify
the whole of the series. Too high a value may result in the algorithm
selecting wrong peaks.

Dimers Allows correct charge assignment for the dimeric component in a

monomer-dimer mixture. In this case, the monomeric series will obscure
alternate peaks in the dimeric series. Therefore, to identify the dimer, the
algorithm must assume a difference of two charge states rather than one
between the two identified peaks.

Threshold… Specifies a minimum intensity of peaks for the algorithm to consider. It is

%BPI specified as a percentage of the intensity of the most intense peak in the
spectrum.

Reject… sd’s A molecular mass is calculated for each peak in the series. The mean
molecular mass and standard deviation of that mean are then calculated.
This parameter offers the opportunity to discard any peak whose molecular
mass is too far from the mean value. Such peaks are discarded and the
mean is recalculated. This feature prevents outlying peaks from biasing
the mean molecular mass measurement. The value is specified as a
number of standard deviations in Da units. The default value of 2.00
means “Reject any peak whose molecular mass lies two or more standard
deviations from the mean”. Two is a safe value, as masses usually will be
within two standard deviations of the mean.

Set this parameter to a high value (e.g. 10.00) if this feature is not required.

Delete Removes the current measured component.

Measure Calculates a component series and displays the result in the Found
component Frame.

Page 7-51
Spectrum

Found Displays the result of the component series calculation.

component
Frame
Molecular The calculated component mass.
Mass:
Missing peaks: The expected masses of peaks that were not found by the calculation.

OK Writes any processing carried out while the dialog is active is to disk. The
component table will be modified and the currently active spectrum
window will be updated to reflect any changes.

Delete All Removes all the components for the particular scan. The components are
not deleted from disk when the OK button is selected.

To Find Components using the Manual Method:

1. Select the Menu Bar Process, Component, Find Manual command; the Manual Find
Components dialog is invoked.

2. Having visually identified a multiply charged peak series, set the Peak 1 m/z and Peak 2 m/z,
either directly, or by using the mouse, see above.

3. Select the Measure button. The Molecular Mass of the component will be displayed in the
Found component frame. Also shown are the expected masses of peaks that were not found
(Missing peaks:).

4. If the identification of the series is satisfactory, proceed to the next one. Otherwise, selecting
the Delete button will remove the component from the component table, and the process may
be repeated.

5. If required, select the Cancel button to abandon the process and exit the dialog box with no
changes to the component table. To clear the component table completely, select the Delete
All button.

6. Select the OK button, to complete the process.

To Find Components when the Mass Range is Known; the Automatic Find Components
Dialog

The automatic method for finding components uses the Automatic Find Components dialog.

Figure 7.42 The Automatic Find Components dialog

Page 7-52
 Spectrum

Series Definition
Frame
Min length… Specifies the minimum number of peaks in the series.
Peaks
Max std dev… Sets an upper limit on the spread of the molecular masses of the peaks in
Da the series. The units are Daltons.

Peak Specifies the tolerance on the position of each peak in the series. It may
window… need to be increased from its default value of 0.5 Da for statistically poor
Da data. Too low a value will result in the algorithm being unable to identify
the whole of the series. Too high a value may result in the algorithm
selecting wrong peaks. The units are Daltons.

Identify Specifies the number of largest peaks to be displayed, smaller peaks will
largest… not be displayed.
single peaks
Allow dimers Allows correct charge assignment for the dimeric component in a
monomer-dimer mixture. In this case, the monomeric series will obscure
alternate peaks in the dimeric series. Therefore, to identify the dimer, the
algorithm must assume a difference of two charge states rather than one
between the two identified peaks.

Peptide filter Specifies that rules for charge assignment will be made; this allows correct
charge assignment for smaller molecules such as peptides.

Mass Range
Frame
Min mol Specifies the lowest molecular mass that the algorithm can consider for a
mass… Da peak series.

Max mol Specifies the highest molecular mass that the algorithm can consider for a
mass… Da peak series.

Find
Components
Frame
Threshold… Specifies the minimum intensity of peaks for the algorithm to consider. It
%BPI is specified as a percentage of the intensity of the most intense peak in the
spectrum.

All Finds all components for the currently active spectrum, according to the
specified parameters.

First Finds only the first component for the currently active spectrum. The
algorithm first considers peaks of highest intensity, then in descending
intensity. After the first component is found and displayed, the First
button changes to Next, allowing the next component to be found.

Page 7-53
Spectrum

OK Writes any processing carried out while the dialog is active is to disk. The
component table will be modified and the currently active spectrum
window will be updated to reflect any changes.

Delete All Removes all the components for the particular scan. The components are
not deleted from disk when the OK button is selected.

To Find Components using the Automatic Method:

1. Select the Menu Bar Process, Component, Find Auto command. The Automatic Find
Components dialog is invoked.

2. Set the Series Definition frame parameters, see above.

3. Set the Mass Range parameters. It is sensible to restrict the range as much as possible; the
wider the mass range the algorithm is allowed to search over, the greater the chance of it
making a series from peaks in the noise.

4. Set the Threshold parameter. A sensible threshold keeps the algorithm out of the noise, and
helps to avoid the above problem.

5. Selecting the First button makes the algorithm find the best series containing the most intense
unassigned peak. If no such series can be identified, then some of the parameters must be
relaxed. First, check the Min length and Threshold parameters. If their values are
reasonable, try larger values for Max std dev and/or Peak window.

6. Selecting the All button causes the algorithm to identify all component series present in the
spectrum subject to the specified parameters.

7. If required, select the Cancel button to abandon the process and exit the dialog box with no
changes to the component table.

8. To clear the component table completely, select the Delete All button.

9. When all components have been identified, select the Close button.

Editing Components for Transform

General

After the components present in the sample have been identified, the Edit Components dialog can
be used to:

• Rename a component.

• Delete a component.

• Sort and re-label the components in order of ascending molecular mass.

• Add a component at known molecular mass, for instance singly-charged species.

• Reject a single peak from the peak series. With poor data, this may improve the accuracy of
the molecular mass.

• Print a report showing all the peaks in the peak series for one or all components.

Page 7-54
 Spectrum

The Edit Components dialog

The Edit Components dialog is invoked by the Menu Bar Process, Component, Edit command.

Figure 7.43 The Edit Components dialog

Add component
Frame
Molecular Specifies the molecular mass.
mass
Add Adds the component mass and label entered in the edit controls to the list
box.

Change label
Frame
Component Enter an identification label for the component (three characters
label maximum).

Update Uses the component label entered in the edit control to update the list box
and the currently active spectrum window.

Component /
charge delimiter
Frame
None When selected, no delimiting character is placed between the component
name and the corresponding charge in the spectral peak annotations.

Page 7-55
Spectrum

Delimiter Allows a delimiting character to be placed between the component name

and the corresponding charge in the spectral peak annotations. For
example, selecting the ‘:’ character would annotate a peak in the style of
Com:12. Any character is valid.

Update Uses the delimiter label entered in the edit control to update the peak
annotations in all spectrum windows.

Edit When selected, the component currently highlighted in the list box is
displayed in the Edit Component dialog which allows editing of the
individual peaks, see the “The Edit Component dialog” section, below.

Sort Sorts the components in the list box by mass and labels will be updated.
The currently active spectrum window will be updated to reflect any
changes.

Delete Deletes the component currently highlighted in the list box from the list
box. The currently active spectrum window is updated to reflect any
changes, however, the component is only deleted from the component
table if the OK button is selected.

Delete All Deletes all components from the list box. The currently active spectrum
window is updated to reflect any changes, however, the component is only
deleted from the component table if the OK button is selected.

Print Prints the currently selected component.

Print All Prints all the components displayed in the list box.

OK When selected, the component table is modified and written to disk. The
currently active spectrum window will be updated to reflect any changes.

The Edit Component dialog

The Edit Component dialog is invoked by the Edit Components dialog, Edit button; it allows
editing of the individual peaks.

Figure 7.44 The Edit Component dialog

A particular component can be rejected, or included, in the series by selecting either the Reject or
Include button respectively.

Note:
Only those peaks lying within the spectrum range when the component was added will be listed.

Page 7-56
 Spectrum

To add a New Component at a Known Molecular Mass

1. Select the Menu Bar Process, Component, Edit command. The Edit Components dialog is
invoked.

2. Enter the component’s mass in the Molecular mass box.

3. Select the Add button. The component is inserted into the component table using the next
available label.

To Change the Name of a Component

1. Select the Menu Bar Process, Component, Edit command. The Edit Components dialog is
invoked.

2. Select the component to be renamed in the list box.

3. Enter the new name for this component (three characters maximum) in the Component Label
text box.

4. Select the Update button.

To Change which Peaks are used in the Calculation of a Component's Molecular Mass

1. Select the Menu Bar Process, Component, Edit command. The Edit Components dialog is
invoked.

2. In the list box, select the component whose peak series is to be to changed.

3. Select the Edit button. The Edit Component dialog is invoked; this displays the peak series
for that component. The peaks that are included in the calculation of the molecular mass of
that component are indicated by a check mark [x].

4. To prevent a peak from being used in the calculation of the component’s molecular mass,
select the peak in the list box, then select the Reject button; the check mark is removed from
the component.

5. To add a peak to the calculation of the component's molecular mass, select the peak in the list
box, then select the Include button.

6. Select the OK button to close the dialog.

To Delete Components

1. Select the Menu Bar Process, Component, Edit command. The Edit Components dialog is
invoked.

2. Select the component to be deleted from the list box.

3. Select the Delete button; select the Delete All button to delete all the components.

To Sort and Re-label the Components

1. Select the Menu Bar Process, Component, Edit command. The Edit Components dialog is
invoked.

2. Select the Sort button. This will sort the components in order of ascending mass and re-label
them, starting at A.

Page 7-57
Spectrum

To Print the Peak Series for a Single Component

1. Select the Menu Bar Process, Component, Edit command. The Edit Components dialog is
invoked.

2. Select the component to be printed from the list box.

3. Select the Print button.

To Print the Peak Series for All Components

1. Select the Menu Bar Process, Component, Edit command. The Edit Components dialog is
invoked.

2. Select the Print All button.

To Use a Component/Charge Delimiter

A delimiter can be used to separate the component label from the charge on m/z spectra.

1. Select the Menu Bar Process, Component, Edit command. The Edit Components dialog is
invoked.

2. Select the Delimiter: option and enter the required delimiter in the adjacent text box.

3. Select the Update button. The spectrum labels are updated to include the delimiter.

The Transform Process

General

When components have been identified in the spectrum, the data system can assign charge states
to each peak. The Transform algorithm uses this information to display the m/z spectrum on a true
molecular mass axis.

To Transform an ElectroSpray Spectrum onto a Molecular Mass Axis; The Transform dialog

The Transform dialog is invoked by the Menu Bar Process, Transform command.

Figure 7.45 The Transform dialog

Min mol mass… Specifies the lowest molecular mass that the algorithm can consider for a
Daltons peak series.

Max mol mass… Specifies the highest molecular mass that the algorithm can consider for a
Daltons peak series.

Resolution… Specifies the resolution (in Da) between data points in the Transformed
Da/channel spectrum.

Page 7-58
 Spectrum

Cut at Frame Allows the User to specify how the m/z spectrum is to be divided up.

Lowest point When selected, regions of equal charge extend to midway between
identified peaks.

Lowest point may produce a superior transform when not all the
components in the spectrum have been identified, or the sample contains
overlapping series.

Mid point When selected, regions of equal charge are divided at the lowest point
between identified peaks.

To carry out a Transform:

1. Identify components in the spectrum as described in the “Finding Components for Transform”
section, on page 7-50.

2. Select the background subtracted continuum spectrum.

3. Select the Menu Bar Process, Transform command; the Transform dialog is invoked.

4. Set the parameters as required, see above.

5. Select the OK button, the Transform process will start.

MaxEnt 1

Introduction

The MaxEnt algorithm uses the maximum entropy method to produce true molecular mass spectra
from multiply-charged ElectroSpray spectra. It has been successfully applied to biopolymers such
as proteins and oligonucleotides. The algorithm has several distinct advantages over the
Transform process:

• MaxEnt automatically finds the molecular weights of the components in a protein mixture
without any knowledge other than that they lie within a specified mass range. This can be
large, e.g. 5 to 100 kDa. To reduce processing time, the technique currently involves a
preliminary survey run, producing a coarse output to find the approximate masses of the
components present.

• The reconstructed MaxEnt spectrum exhibits enhanced resolution and signal-to-noise ratio.

• The reliability of the result can be assessed by probabilistic methods. Thus, a probable error
range can be calculated for each mass.

• MaxEnt data are as quantitative as any ESMS data. The areas under the peaks in the MaxEnt
profile spectrum are representative of the summed intensities of each component's
multiply-charged series in the original m/z data.

Transform works from the raw m/z data, combining the peaks from each component into a single
peak on the molecular mass scale. Because several peaks in the m/z data are used to produce a
single peak in the Transform, the Transformed spectrum exhibits better signal-to-noise than the
raw data. However, the Transformed peaks are no better resolved than in the original data.

MaxEnt retains the mass accuracy given by Transform on components that are adequately resolved
in the original data. In addition, because of its ability to reveal resolution of peaks which is not

Page 7-59
Spectrum

apparent in the raw data, MaxEnt allows the mass measurement of components which were
previously too poorly resolved for mass measurement in the transformed spectrum.

MaxEnt finds the simplest molecular mass spectrum (spectrum of maximum entropy) that could
account for the observed m/z data. The algorithm works iteratively; it takes an initial
approximation to the molecular mass spectrum, and then uses programmed knowledge of
chemistry and mass spectrometer physics (the damage model) to synthesize a corresponding m/z
spectrum (the mock spectrum). It then compares the mock data to the observed (real) data, and
uses the difference between the two to guide it to an improved molecular mass spectrum. The
algorithm terminates when there is sufficiently little difference between mock and real data.

A MaxEnt damage model describes the shape and width of the peaks in the observed m/z data,
which is a composite of two effects. One effect is chemical; the distribution of molecular isotopes
has a characteristic shape that is a function of molecular mass. The other effect is physical, caused
by diffraction effects in the mass spectrometer. The latter effect, alone, can be seen by running a
monoisotopic sample, for instance, Caesium Iodide.

The current implementation of MaxEnt provides a single damage model. This is a Gaussian curve
of constant width, which is a composite model of both of the above effects. To use this model, the
width of a peak in the observed m/z data at half height must be measured.

Note:
The MaxEnt algorithm needs to accurately measure noise within a data file. For this reason, the
Ion Counting Threshold should be set to zero when acquiring data that will be analyzed using
MaxEnt, see the appropriate Instrument User's Guide for further details.

The MaxEnt 1 dialog

The MaxEnt 1 dialog is invoked by the Menu Bar Process, MaxEnt 1 command.

Figure 7.46 The MaxEnt 1 dialog

Page 7-60
 Spectrum

Output Mass
Frame
Ranges Specifies the output mass ranges. A single range is input as two numbers
separated by a colon. Multiple ranges are separated by a comma.

Resolution… Specifies the resolution of the output.

Da/channel

Damage Model
Frame
Uniform Selects the Uniform Gaussian damage model.
Gaussian
Width at half Enter the appropriate value for this parameter in the text box. For a
height… Da detailed discussion on determining the correct value for this parameter, see
the “How to Establish the Correct Peak Width Parameter to Use When
Processing Multiply-Charged Data by MaxEnt” section, on page 7-62.

Simulated Selects the Simulated Isotope Pattern damage model.

Isotope Pattern
Spectrometer Enter the appropriate value for this parameter in the text box.
Blur Width…
Da

Minimum The parameters in this frame place limits on the relative heights of
Intensity Ratios adjacent peaks in the same series
Frame
Left… % Sets the limit for the relative heights of adjacent peaks at the low mass end
of the spectrum. For example, if the parameter is set to 30%, and the most
intense peak in the series is the 15+ peak, the 16+ peak must then be at
least 30% as intense as the 15+ peak, the 17+ peak must be at least 30% as
intense as the 16+ peak, etc.

Right… % Sets the limit for the relative heights of adjacent peaks at the high mass
end of the spectrum. For example, if the parameter is set to 40%, then the
14+ peak must be at least 40% as intense as the 15+ peak, the 13+ at least
40% as intense as the 14+, etc.

Note:
The default values of 33% for each of these parameters will always work,
but, for most data sets, these values can profitably be increased. In
particular, when doing a survey run, increasing the Left and Right
Minimum Intensity Ratios will give significant reduction in the intensity
of the “harmonic artifacts”, i.e. the peaks at twice, three times, etc. the
mass of each component.

Completion
options Frame
Iterate to When selected, the MaxEnt process will continue iterating until it
convergence converges.

Page 7-61
Spectrum

Maximum When selected, the MaxEnt process will perform up to the number of
number of iterations specified in the adjacent text box.
iterations
Exit dialog on When selected, MaxEnt will automatically accept the results, exit the
completion MaxEnt dialog, and display the MaxEnt spectrum on completion. If this
option is not selected, the MaxEnt dialog will remain displayed on
completion, giving the User the option to accept the results and save the
MaxEnt spectrum, or discard the results.

OK Starts the MaxEnt process.

To Produce a Survey Spectrum

The sole purpose of producing a survey spectrum is to determine the approximate masses of the
components present. It is possible to analyze a complete unknown by selecting a very wide output
mass range, e.g. 10 to 100 kDa. Usually, the major components are revealed after three or four
iterations.

1. Background subtract the data, refer to the “The Background Subtract Process” section, on
page 7-40. In the Background Subtract dialog (see Figure 7.33, on page 7-41), set the
parameters to fit an appropriate polynomial with 30 to 50% of the data below it. 30% usually
leaves a low level of noise in the MaxEnt result; the User may wish to increase this for noisier
spectra.

2. MaxEnt will process the data actually on display. Hence, the User can “rubber-band” the
display to exclude parts of the spectrum that contain noise. This can improve the MaxEnt
result in some cases. In addition, if the spectrum has a flat baseline, it is possible to remove
this with the mouse, “rubber-banding” in the vertical direction.

3. Select the Menu Bar Process, MaxEnt 1 command, the MaxEnt 1 dialog is invoked, see the
“The MaxEnt 1 dialog” section, on page 7-60.

4. Set up the Output Mass range, in the Ranges text box. The mass range is given as two
numbers separated by a colon, e.g. 10000:100000.

5. The Resolution parameter controls the “texture” of the result. Set this parameter to a value in
the range 10 to 25 Da/channel. This will give a coarse result, not showing fine detail and
without accurate masses, but the spectrum will suffice to locate the major components for a
finer run over a smaller mass range.

6. Select the required damage model option in the Damage model frame. To use the Uniform
Gaussian model, the average width at half height of a peak in the m/z spectrum must be
estimated. For a detailed discussion on determining the correct value for the Width at half
height parameter, see the “How to Establish the Correct Peak Width Parameter to Use When
Processing Multiply-Charged Data by MaxEnt” section, below.

7. Set the Left and Right Minimum Intensity Ratio parameters.

8. Select the required options in the Completion options frame.

9. Select the OK button. The MaxEnt status dialog will appear. The algorithm will initialize
itself, then draw molecular mass axes, and the first iteration will start.

How to Establish the Correct Peak Width Parameter to Use When Processing
Multiply-Charged Data by MaxEnt

When processing data by MaxEnt, it is crucial that the correct peak width at half height is used.
The only sure way to establish this width is to measure it, using peaks that are known to be
singlets.

Page 7-62
 Spectrum

The ideal way is to measure the width of a known singlet in the m/z spectrum to be processed. For
example, in a haemoglobin spectrum, it may be required to separate and measure the components
in an unresolved β-globin doublet, when it is known that the α-globin is a singlet. The measured
width of an α-globin peak near the center of the spectrum may then be used directly in the MaxEnt
processing, since the molecular weights of the two globins are similar.

In many situations, however, the peaks in the sample data will not be sufficiently resolved for their
widths at half height to be measured. In these cases, it is necessary to measure the peak width
from a multiply-charged spectrum run under identical conditions as the sample spectrum, and
known to contain singlets. This can be the spectrum used to calibrate the sample spectrum, or
another spectrum containing known singlets. In either case, it will generally be necessary to
correct the measured peak width in order to find the value to use when processing the data by
MaxEnt. This is derived as follows:

Let the measured width at half height of a singlet in the 'calibration' spectrum be wc and let the
peak have nc charges.

Let the molecular weights of the ‘calibration’ compound and the sample be Mc and Ms
respectively.

Let the theoretical widths at half height due to the isotopic distribution of the elements in the
molecule be Wc and Ws for Mc and Ms respectively. These may be found from Figure 7.47.

It is assumed that the width of a peak in the m/z spectrum is made up of two components; a
component due to the theoretical isotopic distribution and a component due to the instrument itself
(wi). These are assumed to be Gaussian, and are added as the root of the sum of the squares.

Hence,

wc2 = wi2 + (wc/nc)2 - - - - - - - - - - - - (1)

and

ws2 = wi2 + (Ws/ns)2 - - - - - - - - - - - - (2)

where ws is the width required for processing the sample spectrum by MaxEnt, and ns is the
number of charges on a peak at a similar part of the m/z spectrum to that used for measuring wc.

Combining (1) and (2) to eliminate wi,

ws2 = wc2 + (Ws/ns) 2 - (Wc/nc)2 - - - - - (3)

Example

Suppose using myoglobin (Mr = 16951.5), wc was measured as 1.0 Da for the m/z 1212 peak
(nc = 14). From the graph, Wc = 8.2 Da.

Suppose, also, that the sample has a molecular weight of ~40000 Da. From the graph,
Ws = 12.6 Da. At m/z ~1212, ns = 33.

Using equation (3),

ws2 = 1.0 + (12.6/33)2 - (8.2/14)2 = 0.80

or ws = 0.90.

If wc = 0.8 for myoglobin, ws = 0.67 for the 40 kDa protein.

Page 7-63
Spectrum

160
15 0
1 40
1 30
120
11 0
100
M olecular W eight (kD a)
90
80
70
60
50
40
30
20
10
0
30

25

20

H eight 1 5

10

0
at Half
W idth
Peak

(D a)

Figure 7.47 Theoretical Peak Width of Proteins due to Isotopic Distribution vs. Molecular
Weight

Page 7-64
 Spectrum

Interpreting the Survey Spectrum

Figure 7.48 shows the first three iterations of a MaxEnt survey run on a data set produced from
leech haemoglobin.

After the first iteration, the major components are already visible, but the harmonic artifacts at
twice the mass of each component are present at significant intensity. Also, the background level
is high, and rises with increasing mass. After the second iteration, the intensity of artifacts and
background level has been greatly reduced. Neither is present with significant intensity after three
iterations.

Sub-harmonic artifacts at fractions (half, quarter, etc.) of the true molecular mass for the first
couple of iterations may also be seen.

Figure 7.48 First three iterations of a MaxEnt survey run on leech haemoglobin

To Stop a MaxEnt Run Before the Algorithm Converges

1. Select the Halt button.

2. The result may now be accepted by selecting the OK button, or discarded by selecting
Cancel. MaxEnt may also be restarted by selecting the Restart button.

3. If the spectrum is accepted and MaxEnt is to be started later, this may be done by selecting the
Menu Bar Process, MaxEnt command again.

To Produce the Definitive MaxEnt Spectrum

Once the approximate masses of the major components are known, whether from prior knowledge
of the sample, or a MaxEnt survey run, the definitive MaxEnt spectrum revealing all the fine
structure can be produced:

1. Either select the background subtracted data used to produce the survey spectrum, or use
Background Subtract to produce some.

2. Select the Menu Bar Process, MaxEnt 1 command; the MaxEnt 1 dialog is invoked.

Page 7-65
Spectrum

3. Set up the Output Mass range from the knowledge of the approximate masses of the major
components. The run time of MaxEnt is directly proportional to the number of data points in
the output, and this number is the product of mass range and reciprocal of resolution.
Therefore, do not set the mass range unnecessarily wide.

4. Note that two, or more, Output Mass Ranges separated by commas may be selected, e.g.
16500:17500, 24500:26500. Using this facility reduces the processing time. The Output
Mass Ranges should include all the significant components found in the survey run, in order
to make the definitive MaxEnt spectrum a faithful representation of the original data.

5. Set the Resolution parameter to 1.0 Da/channel. Generally, this is sufficiently small to ensure
there will be several data points across each peak in the output, and a centroid can be taken to
give an accurate mass. Occasionally, a smaller value e.g. 0.5 Da/channel may be necessary.
This will increase the processing time, however.

6. Set the Damage model and Minimum intensity ratios parameters, as described above.

7. Select the OK button.

Mock Data

To get a definitive result, MaxEnt must to run to completion. It will then produce two spectra; one
is the MaxEnt result on a molecular mass axis, and the other is the mock data, explained above.
Examining the mock data can help decide how good the parameter settings were. Mock data
should fit the observed data within the tolerance of the noise.

Figure 7.49 shows mock data (upper) and original data (lower).

Figure 7.49 Original data from leech haemoglobin (lower), and MaxEnt mock data (upper)

To Examine the Fit of Mock to Real Data

1. Select the Menu Bar Display, View command. The Spectrum Display dialog is invoked, see
the “Controlling the Appearance of the Display” section, on page 7-28.

2. Select the Style frame, Overlay Graphs option. This will cause spectra in the same window
to be superimposed.

Page 7-66
 Spectrum

3. Select the Normalize Data to: frame, Link Vertical Axes option.

4. Select the OK button.

5. The raw data must now be displayed in the window containing the mock data. Click inside
the window containing the mock data, then select the Menu Bar File, Open command. The
Spectrum Data Browser dialog is invoked.

6. Ensure that the Add Data radio button is selected, then select the raw data from the list box.

7. Select the OK button.

The Minimum Intensity Ratio parameters will affect the intensities of the peaks in the mock data,
and the appropriate damage model width parameter will affect the widths of the peaks in the mock
data.

Mass Measurement of MaxEnt Spectra

Special interpretation must be placed on peaks in MaxEnt spectra. The topmost point of the peak
is not the most probable estimate of the peak’s mass; rather, a centroid must be taken. The height
of a MaxEnt peak is an indicator of how good an estimate the algorithm can make of the mass.
This means the height is not proportional to the relative concentration of that component in the
sample; but the area is.

There are two ways to produce MaxEnt spectra with accurate masses. The first presents the profile
spectrum labeled with accurate mass values, as in Figure 7.50 (upper). The second presents the
spectrum as bars, with the height of each bar being proportional to the area of the peak in the
profile data, as in Figure 7.50 (lower). Note that the apparent ratio of the intensities of the α and
α1 peaks has changed. The ratios observed in the lower spectrum are definitive, provided Areas
are used.

Figure 7.50 MaxEnt results from leech haemoglobin

Page 7-67
Spectrum

To Produce a Profile Spectrum with Accurate Masses

1. Select the MaxEnt spectrum.

2. Select the Menu Bar Process, Center command. The Spectrum Center dialog is invoked,
see the “To Smooth a Continuum Spectrum” section, on page 7-42.

3. Set the Min peak width at half height (channels) parameter to 1. This will interpret the
smallest, narrowest feature in the spectrum as a peak. If this does not produce the required
result, the value of this parameter can be increased to group the narrower features together
with the wider ones.

4. Select a Center method. Top is provided mainly for compatibility with the LAB-BASE data
system. Centroid top (%) is the recommended method, since the parameter can be set to use
the well-resolved part of the peak only, keeping clear of baseline effects. Recommended
values for Centroid top(%) are in the range 70 to 90%.

5. Ensure the Centered spectrum frame, Create centered spectrum option is not selected.

6. Select the OK button.

To Produce a Bar Spectrum with Heights Proportional to Component Concentration

1. Select the MaxEnt spectrum.

2. Select the Menu Bar Process, Center command. The Spectrum Center dialog is invoked.

3. Set the Min peak width at half height parameter as described above.

4. Select a center method as described above, e.g. Centroid top (%)=90.

5. Select the Centered spectrum frame, Create centered spectrum option.

6. Select the Centered spectrum frame, Areas option.

7. Select the OK button.

MaxEnt Errors

General

A probable error range can be calculated for the mass of each peak in the MaxEnt spectrum.

This is done by sampling the distribution of possible spectra at about a dozen points near the most
probable spectrum. Hence, the error analysis requires a further dozen iterations of the MaxEnt
kernel, and for this reason, it is a separate process.

To Calculate the MaxEnt Errors

1. Form a MaxEnt profile spectrum with accurate masses as described above.

2. Select the Menu Bar Process, MaxEnt errors command. The status dialog will appear, and
the first cloud sample will commence. Twelve samples are performed in all, and after the last
one, the spectrum is redisplayed with the errors.

3. Save the errors by selecting the Menu Bar File, Save command.

Page 7-68
 Spectrum

Note:
The MaxEnt errors will only be seen when the Mass Error parameter has been selected in the
Spectrum Peak Annotation dialog, see the “Controlling the Appearance of Peak Labels” section,
on page 7-31.

MaxEnt Initialization Errors

Occasionally, an error message is displayed when MaxEnt is started: MaxEnt initialisation error
–1, or MaxEnt initialisation error -2. These errors mean that there is not enough memory
available to execute the current MaxEnt operation. In this case:

1. Close down MassLynx and any other Windows programs in order to free all available
memory.

2. Run MassLynx again.

3. Load the spectrum to be processed.

4. Try to run MaxEnt again.

5. If the same error occurs, alter the MaxEnt parameters so that less memory is required.
Reducing the mass range in the Ranges parameter, or increasing the Resolution parameter
can do this.

MaxEnt 2

General

MaxEnt 2 can be applied to any singly charged continuum spectrum to increase resolution and
remove noise. For information about how the MaxEnt process works see the “MaxEnt 1” section,
on page 7-59.

The MaxEnt 2 dialog

The MaxEnt 2 dialog is invoked by the Menu Bar Process, MaxEnt 2 command.

Figure 7.51 The MaxEnt 2 dialog

Peak Width at Enter the appropriate value for this parameter in the text box.
Half Height…
Da
Tolerance Increasing this parameter makes the algorithm terminate sooner, but the
result may be inferior. The normal operating range for this parameter is
between 0.030 and 0.300.

Rate Controls the rate at which the MaxEnt Reconstruction process will run. It
is recommended that values between 1.00 and 3.00 be used for this
parameter.

Page 7-69
Spectrum

Overlay When selected, the new data calculated with each MaxEnt iteration is
Iterations overlaid on top of the previous data.

OK Starts the MaxEnt process.

To Use MaxEnt 2

1. Display the singly charged continuum spectrum.

2. Adjust the display range to show the mass Range to be analyzed. MaxEnt will process the
data range which is actually on display, this means that the display can be set to exclude parts
of the spectrum which contain uninterpretable noise.

3. Select the Menu Bar Process, MaxEnt 2 command, the MaxEnt 2 dialog is invoked.

4. Set the parameters as required.

5. Select OK to start the analysis. The MaxEnt Reconstruction status dialog will appear. The
algorithm will initialize itself, then draw molecular mass axes, and the first iteration will start.
The status dialog shows the data produced by each iteration of MaxEnt.

Figure 7.52 The MaxEnt Reconstruction status dialog

6. When the MaxEnt reconstruction has finished the status dialog will display a message that the
algorithm has converged. Select OK to accept the spectrum. MaxEnt will then produce two
spectra; one is the MaxEnt result on a molecular mass axis, and the other is the mock data.

Figure 7.53 shows part of a spectrum obtained from MALDI analysis of a peptide mixture. The
upper trace shows background subtracted raw data, the middle trace shows background subtracted
and smoothed raw data and the lower trace shows the MaxEnt reconstructed data. The MaxEnt
reconstructed data shows much improved resolution of the isotope peaks.

All MaxEnt processed spectra are stored to disk, with the raw data file; they can be selected using
the Spectrum Data Browser, History button.

Page 7-70
 Spectrum

Figure 7.53 Original data from Peptide mixture (upper and middle) and MaxEnt 2 data
(lower)

To Stop a MaxEnt Run Before the Algorithm Converges

1. Select the Halt button.

2. The result may now be accepted by selecting the OK button, or discarded by selecting
Cancel. MaxEnt may also be restarted by selecting the Restart button.

3. If the spectrum is accepted and MaxEnt is to be restarted later, this may be done by selecting
the Menu Bar Process, MaxEnt command again.

MaxEnt 3

General

MaxEnt 3 can be applied to any low mass, multiply-charged continuum spectrum to resolve the
multiply-charged peaks onto a singly-charged axis. The MaxEnt 3 program interprets isotope
clusters to gain charge state information. For more information about how the MaxEnt process
works see the “MaxEnt 1” section, on page 7-59.

The MaxEnt 3 dialog

General

The MaxEnt 3 dialog is invoked by the Menu Bar Process, MaxEnt 3 command.

Note:
The MaxEnt 3 dialog format depends on the type of data selected.

Page 7-71
Spectrum

The MaxEnt 3 dialog (Tof Data)

Figure 7.54 The MaxEnt 3 dialog (TOF data)

Min molecular Specifies the minimum mass for the output mass range.
mass (Da)
Max molecular Specifies the maximum mass for the output mass range.
mass (Da)
Maximum no of Defines the maximum charge state that MaxEnt can identify. For
charges example, a value of 2 means that singly and doubly-charged peaks will be
detected, 3 means singly, doubly and triply charged peaks will be detected,
etc. Do not set this parameter to a higher value than needed, or artifacts
may result.

Peak width
Frame
Peak width 1 Specifies the peak width at half height for the minimum mass specified in
(Da)… at m/z 1 the at m/z 1 text box.

Peak width 2 Specifies the peak width at half height for the maximum mass specified in
(Da)… at m/z 2 the at m/z 2 text box.

Auto peak Determines peak widths automatically.

width
determination Note:
The preceding Peak width options are disabled when this option is
selected.

OK Starts the MaxEnt 3 process.

Advanced Invokes the MaxEnt 3 Advanced Parameters dialog, see the “The
MaxEnt 3 Advanced Parameters Dialog” section, on page 7-73.

Page 7-72
 Spectrum

The MaxEnt 3 dialog (Quad Data)

Figure 7.55 The MaxEnt 3 dialog (Quad data)

Min molecular Specifies the minimum mass for the output mass range.
mass (Da)
Max molecular Specifies the maximum mass for the output mass range.
mass (Da)
Maximum no of Defines the maximum charge state that MaxEnt can identify. For
charges example, a value of 2 means that singly and doubly-charged peaks will be
detected, 3 means singly, doubly and triply charged peaks will be detected,
etc. Do not set this parameter to a higher value than needed, or artifacts
may result.

Peak width (Da) Specifies the peak width at half height.

Advanced Invokes the MaxEnt 3 Advanced Parameters dialog, see the “The
MaxEnt 3 Advanced Parameters Dialog” section, below.

The MaxEnt 3 Advanced Parameters Dialog

The MaxEnt 3 Advanced Parameters dialog is invoked by the MaxEnt 3 dialog Advanced
button.

Figure 7.56 The MaxEnt 3 Advanced Parameters dialog

No. of Ensemble Specifies the number of ensemble members. The recommended setting is
Members 1.

Iterations per Specifies the number of Iterations per Ensemble Member. The
Ensemble recommended setting is 10, or 20.
Member
Compress Data Compresses the spectrum to half its original size, thus reducing run time.
No deterioration in results has been observed as a result of using this
option.

MaxEnt 3 uses an ensemble of processes, notionally working in parallel. Increasing the No of

Ensemble Members may improve results, but will increase run time. The Iterations per
ensemble member parameter is a guide to the amount of CPU time each ensemble member is
allowed. Again, increasing this parameter may result in improved results at the expense of
runtime.

Page 7-73
Spectrum

To Use MaxEnt 3

1. Display the multiply charged continuum spectrum. MaxEnt will process the data range on
display, and the display can be “rubber-banded” vertically to set a noise level.

2. Adjust the display range to show the mass Range to be analyzed. MaxEnt will process the
data range which is actually on display; this means that the display can be set to exclude parts
of the spectrum which contain uninterpretable noise.

3. Select the Menu Bar Process, MaxEnt 3 command; the MaxEnt 3 dialog is invoked.

4. Set the parameters as required.

5. Select the Advanced button to set the Ensemble parameters and the data compression option
in the MaxEnt 3 Advanced Parameters dialog.

6. Select OK to start the analysis. The MaxEnt 3 status dialog will appear, showing the progress
of the process. Select the Cancel button to stop a MaxEnt 3 run before the end of processing.

Figure 7.57 The MaxEnt3 Sequence status dialog

7. When the MaxEnt 3 has finished, MaxEnt will produce two spectra; one is the MaxEnt result
on a molecular mass axis, and the other is the mock data.

Figure 7.58 shows part of a spectrum obtained from TOF analysis of Glu-fibrinopeptide. The
lower trace shows raw data, and the upper trace shows the MaxEnt 3 data. The MaxEnt data
shows the charge state of the isotope peaks interpreted correctly.

Figure 7.58 Original data from Glu-fibrinopeptide (lower) and MaxEnt 3 data (upper)

Page 7-74
 Spectrum

All MaxEnt processed spectra are stored to disk, with the raw data file, and can be selected using
the Spectrum Data Browser, History button.

To Stop a MaxEnt 3 Run Before the Algorithm Converges

1. Select the Cancel button

2. The result may now be accepted by selecting the OK button, or discarded by selecting
Cancel. MaxEnt may also be restarted by selecting the Restart button.

3. If the spectrum is accepted and MaxEnt is to be started later, this may be done by selecting the
Menu Bar Process, MaxEnt command again.

Isotope Cluster Abundance Plots

General

MassLynx can produce an isotope cluster abundance plot for a given formula. For example,
Figure 7.59 shows the predicted isotope model for the formula C9H10N2O2Cl2 (Linuron).

Figure 7.59 Typical Isotope modeling display

The Isotope modelling Dialog

The Isotope modelling dialog is invoked by the Menu Bar Tools, Isotope Model command, or
the Elemental Composition Window Menu Bar Process, Set Isotope Match Parameters
command, see the “The Elemental Composition Window Menu Bar” section, on page 7-82.

Page 7-75
Spectrum

Figure 7.60 The Isotope modelling dialog

Formula Enter the chemical formula for the compound, using standard IUPAC
(International Union of Pure and Applied Chemistry) notation.

A formula can also be transferred from the BioLynx Protein or Nucleic

Acid Editors by selecting the Elemental Composition Window Tool Bar
button to transfer the molecular formula to the Windows Clipboard.
The formula can then be pasted into the Isotope modelling dialog
Formula text box by selecting the Paste button.

Spectrum
Modelling Frame
Create Creates a continuum spectrum as well as the centroided spectrum.
Continuum
spectrum
Peak width at Enter a value for the continuum spectrum.
half-height
Create When selected, the modeled spectrum will contain the multiply charged
ElectroSpray series that are present in ElectroSpray spectra.
spectrum

Isotope cluster
parameters
Frame
Separation Determines the resolution of the modeled spectrum. Peaks that would be
closer together than this value are combined into a single peak.

Min Determines the threshold below which peaks are not considered
Abundance significant.
(%)

Page 7-76
 Spectrum

Molecular Mass This frame contains parameters that determine the range used for the peaks
Range Frame to be displayed in the modeled spectrum.

Full Range Selects the full range.

Mass From Enter a value for the lower end of the range.

Mass To Enter a value for the upper end of the range.

Multiply-
charged ion
Frame
Multiply- Creates a single multiply-charged peak with the number of charges
charged ion specified in the Charge state text box.

OK Updates the isotope structure and initializes the isotope modeling

algorithm.

Paste Pastes a formula from the Windows clipboard into the Formula text box.

Add Adds the spectrum to the current Spectrum Window.

Replace The spectrum replaces the currently selected spectrum in the Spectrum
Window.

New window Displays the spectrum in a new Window.

User elements Invokes the User-definable elements dialog, see the “The User-definable
elements Dialog” section, below.

Print masses Prints out the current range of masses.

The User-definable elements Dialog

The User-definable elements dialog is invoked by the Isotope modelling dialog, User elements
button. It allows the User to specify a list of elements, isotopes, and/or molecules.

Figure 7.61 The User-definable elements dialog

Name Enter a name for the element in this text box.

Page 7-77
Spectrum

Symbol Enter a symbol for the element in this text box.

Mass Enter a mass for the element in this text box.

OK Closes the dialog and returns to the Isotope modelling dialog.

Add Adds the current group to the list. A maximum of ten groups may be
added.

Deletes Deletes the currently selected group from the list.

Update Updates the currently selected group with changed details from the Name,
Symbol, or Mass text boxes.

To Produce an Isotope Cluster Abundance Plot

1. Select the Spectrum Menu Bar Tools, Isotope Model command, or the Elemental
Composition Window Menu Bar Process, Set Isotope Match Parameters command; the
Isotope modelling dialog is invoked.

2. Set the parameters as required, see above.

3. Select the OK button.

Elemental Composition
General

Elemental Composition processes either a single or a multiple number of molecular masses, such
that the elemental composition of the molecule(s) may be estimated.

The EleComp Parameters Dialog

The EleComp Parameters dialog is invoked by the Menu Bar Tools, Elemental Composition
command.

Figure 7.62 The EleComp Parameters dialog

Use Single Mass Select a single mass by entering the value in the text box, or by
right-clicking on a peak in the display.

Use Multiple Selects the set of masses currently displayed.

Masses
OK Invokes the Elemental Composition dialog and an elemental composition
report is generated from the selected peak(s).

Page 7-78
 Spectrum

To Produce an Elemental Composition Report

1. Select the Menu Bar Tools, Elemental Composition command; the EleComp Parameters
dialog is invoked.

2. Set the parameters as required, see above.

3. Select the OK button; an Elemental Composition Report is created and displayed in the
Elemental Composition window, see the “The Elemental Composition Window” section, on
page 7-80.

When the Elemental Composition Window is displayed, there are two other ways of generating
reports:

• A Spectrum list can be copied from another Windows application and pasted into the
Elemental Composition window by selecting the Tool Bar button, or selecting the Menu
Bar Edit, Paste command. The software will automatically generate a new report.

• Selecting the Elemental Composition window Tool Bar button, or selecting the Menu
Bar Process, Enter Single Mass command will invoke the Mass dialog.

Figure 7.63 The Mass dialog

Enter a new Mass, or select a previously entered mass from the drop down list box. Select the
Display all option to display all the results found for a mass. To display a limited number of
results per mass, deselect the Display all option and select the arrows to change the value
in the number box. For example, if 5 is entered, the five closest results to the mass will be
displayed.

If the Match isotopes option is selected, the Edit Parameters button is enabled; this invokes
the Isotope Cluster Parameters dialog. Enter values for the Separation and Min.
Abundance (%).

Note:
The Isotope Cluster Parameters dialog may also be invoked by the Elemental Composition
window Menu Bar Process, Set Isotope Cluster Parameters command.

Page 7-79
Spectrum

Figure 7.64 The Isotope Cluster Parameters dialog

Select the Mass dialog OK button to generate the Elemental Composition Report.

To Update an Elemental Composition Report

The search details for a report can be changed by selecting the Elemental Composition window
Tool Bar button, or by selecting the Menu Bar Process, Set Parameters option. This
invokes the Parameters dialog; change the required details and select the OK button. For more
information on the Parameters dialog, see the “Elemental Composition Parameters” section, on
page 7-84.

The Elemental Composition Window

The Elemental Composition window is invoked, via the EleComp Parameters dialog, from the
Spectrum Menu Bar Tools, Elemental Composition command, see the “To Produce an Elemental
Composition Report”, on page 7-79.

Figure 7.65 Typical Elemental Composition Window

Page 7-80
 Spectrum

The window is split into three panes:

• The top pane shows details of the number of masses used (if there are more than one) and the
number of possible compounds found.

• The bottom pane displays the spectrum of the components found.

• The middle pane shows the following details for the compounds found:

Mass The mass being analyzed. Note that for multiple masses, each mass will
have a set of results.

RA The percentage relative abundance; this is determined by expressing the

intensity of the mass as a percentage of the intensity of the most intense
peak. This is only displayed for multiple mass calculations.

Calc Mass The calculated mass for the formula shown in the Formula column,
subject to the specified tolerance limits.

mDa The difference between the calculated mass and the entered mass, in
milliDaltons.

PPM The difference between the calculated mass and the entered mass, in parts
per million.

DBE The double bond equivalent for the formula shown in the Formula
column.

Formula A suggested formula for the entered mass. The formula is not a true
atomic formula; it is merely a summation of the quantities of elements,
isotopes, and/or superatoms that may compose the sample. In the formula,
each symbol is displayed followed by the number of times of its
occurrence.

Score The Score relates to the fit of the reconstructed isotopic spectrum for each
hit, to the original spectrum. The score is given as a rank, with a rank of 1
corresponding to the best match to the original spectrum. If the rank is
given as “n/a”, the reconstructed isotopic spectrum contains no masses in
the mass range of the original spectrum.

Symbol A symbol column is created in the middle pane for each

columns element/isotope/superatom included in determination of possible elemental
compositions. The value that appears in the symbol column on any given
row is the number of occurrences of that element in the corresponding
elemental composition as shown in the Formula column.

Clicking on the Mass column heading will list the masses in reverse order. Clicking on any of the
other column headings will display the values in ascending order for each mass, clicking a second
time will display them in descending order for each mass.

Holding the mouse cursor over the RA, DBE and symbol column headings will display the
minimum and maximum values defined on the Parameters dialog. Holding the mouse pointer
over the mDa and PPM column headings will display the tolerance values defined on the
Parameters dialog.

A status bar is located at the foot of the main application window. When masses are not currently
being processed, the text aligned to the left of the bar simply says “For Help, Press F1”. During
processing, it displays processing information; for many of the larger spectra, the calculation may
take several minutes to complete.

Page 7-81
Spectrum

A sunken pane, to the right of the progress text, displays the last mass that is to be processed. It is
impossible to determine in advance how long the process will take, due to the nature of the
formulae calculations. Therefore, a progress bar cannot be provided. However, by displaying the
final mass to be processed, the User can determine approximately how long the calculations will
take and therefore make the decision as to whether to terminate the process, if required.

The Elemental Composition Window Menu Bar

The Elemental Composition Window File Menu

Figure 7.66 The Elemental Composition Window File Menu

Save Results Saves the results in a plain text file (*.txt). A standard Save As dialog is
invoked.

Load Settings Loads a previously saved Parameter file. Select the required *.els file
from the invoked Load Settings dialog. The software will automatically
generate a report using these settings.

Save Settings Saves a Parameter file. Enter a name in the invoked Save Settings dialog
and select the Save button.

Print Results Sends the table of results to the printer.

Print Preview Standard Windows command.

Print Setup Standard Windows command.

Exit Closes the Elemental Composition Window.

The Elemental Composition Window Edit Menu

Figure 7.67 The Elemental Composition Window Edit Menu

Copy If the top or middle pane is active the current set of results is copied to the
clipboard. The format is copied as plain text and formatted in columns in
the exactly the same manner as it is saved (see above). If the bottom pane
is active, the spectrum on display is copied to the clipboard.

Page 7-82
 Spectrum

Paste Pastes a mass spectrum that has been copied to the clipboard, into the
application. The application then processes the data and generates the
results, which are displayed in the results table.

This button is disabled if there is nothing on the clipboard. If the clipboard

data isn’t useable spectrum data, no processing is carried out.

Clear Results Deletes the current set of results in the table and resets the information
window.

The Elemental Composition Window View Menu

Figure 7.68 The Elemental Composition Window View Menu

Toolbar Toggles the Tool Bar on and off.

Status Bar Toggles the Status Bar on and off.

Formula Toggles the Formula column on and off.

Table Toggles the symbol columns on and off.

The Elemental Composition Window Process Menu

Figure 7.69 The Elemental Composition Window Process Menu

Enter Single Invokes the Mass dialog; see the “To Produce an Elemental Composition
Mass Report” section, on page 7-79.

Set Parameters Invokes the Parameters dialog; see the “Elemental Composition
Parameters” section, on page 7-84.

Set Isotope Invokes the Isotope Cluster Parameters dialog; see the “To Produce an
Cluster Elemental Composition Report” section, on page 7-79.
Parameters
The Elemental Composition Window Help Menu

Figure 7.70 The Elemental Composition Window Help Menu

Help Topics Invokes the MassLynx Help function.

About Elemental Displays information about Elemental Composition.

Composition

Page 7-83
Spectrum

The Elemental Composition Window Tool Bar

Tool Menu equivalent Purpose

Bar
button
File, Save Results Saves the Elemental Composition Report.

Edit, Copy Copies the selection onto the clipboard.

Edit, Paste Pastes the contents of the clipboard into the Elemental
Composition Window.

Edit, Clear Results Clears the results from the Elemental Composition
Window.

File, Print Prints the Elemental Composition Report.

Process, Enter Single Invokes the Mass dialog.

Mass
Process, Set Parameters Invokes the Parameters dialog.

Process, Set Isotope Invokes the Isotope modelling dialog.

Match Parameters
Resets the Spectrum display to its default magnification.

Elemental Composition Parameters

General

The Parameters dialog is invoked by selecting the Elemental Composition Window Tool Bar
button, or selecting the Menu Bar Process, Set Parameters option. The dialog has two
pages:

• General Parameters.

• Symbol Parameters.

Selecting the Reset to Defaults button will set the Hot Symbols (see the “Specifying Hot
Symbols” section, on page 7-88) and all other fields on the Parameters dialog to the default
values.

Page 7-84
 Spectrum

The Parameters Dialog; General Parameters Page

Figure 7.71 The Parameters dialog: General Parameters page

Tolerance Frame

MilliDaltons: Expresses the parameters that comprise the tolerances for subsequent
calculations as milliDaltons (mDa).

PPM: Expresses the parameters that comprise the tolerances for subsequent
calculations as parts per million (PPM).

Note:
1. The above two options express the same thing. However, it is
sometimes more convenient to use one rather than the other. Both are
used to indicate how large a tolerance to allow when the cumulative
mass of any given composition formula has been calculated.

2. The calculations use plus and minus this value, so the default
200 mDa and 5 PPM represent ± 200 mDa and ± 5 PPM.

Minimum % Specifies the minimum relative abundance, i.e. specifies that only masses
RA: with a relative abundance above the specified value are to be processed.

Page 7-85
Spectrum

Results Frame The Results controls specify the quantity of results to display. They are
useful for reducing the time it takes to tabulate the results immediately
following calculation.

Display only Causes only those masses that fit within the specified tolerance parameters
valid results to be displayed. Otherwise, invalid masses are also listed with “-” in the
various columns to indicate that they have no results.

No. Results to When selected, up to 1000 results will be displayed for each mass, if they
display: comply with the specified tolerance parameters. If this option is not
selected, the number of results displayed will be no greater than the value
specified in the adjacent text box.

Double Bond The Double Bond Equivalent (DBE) specifies the Maximum: and
Equivalent Minimum: number of double bonds per molecule considered to be
Frame acceptable in producing a valid formula. A simple calculation can be
performed which will indicate how many double bonds there would be
within a molecule of any given formula. The result of this calculation is
then used to determine whether a given result falls within the specified
validation limits.

Mass Mode The mass modes utilize the Monoisotopic, Chemical, or Nominal mass of
Frame the active symbols when performing the calculations.

Electron State The Electron State specifies the preferred types of molecule to be
Frame included or excluded from the results.

Both odd and Includes both odd and even electron types.
even
Odd electron Includes odd electron types only.
only
Even electron Includes even electron types only.
only

Page 7-86
 Spectrum

The Parameters Dialog; Symbol Parameters Page

General

Figure 7.72 The Parameters dialog: Symbol Parameters page

Element Limits Up to ten “Hot Symbols” may be specified in this frame, to allow their
Frame properties to be changed quickly and easily. Any element, isotope, or
superatom can be specified as a Hot Symbol as shown in the “Specifying
Hot Symbols” section, on page 7-88.

Check box Selects a Hot Symbol.

Symbol button Invokes the Select Hot Symbol dialog, see the “Specifying Hot Symbols”
section, on page 7-88.

From The minimum number of elements or isotopes that the calculated formula
must contain; e.g. if the From value for Cl is 2, the formula must contain
Cl2, but can contain any number above this, e.g. Cl3, Cl4, etc.

To The To value is the maximum number of elements or isotopes that the

calculated formula must contain; e.g. if the To value for Cl is 2, the
formula must contain Cl2, but can contain any number below this.

Note:
If the From and To values are the same, the calculated formula must
contain this exact number of elements or isotopes; e.g. if the From and To
values for Cl are both 2, then the formula must contain Cl2.

Periodic Table Invokes the Periodic Table dialog, see the “To Select an Element” section,
on page 7-89.

Page 7-87
Spectrum

Deselect All Deselects all the Hot Symbols.

Superatoms
Frame
Tables Invokes the Superatom Tables dialog, see the “Superatom Tables”
section, on page 7-91.

Limits Invokes the Superatom Limits dialog, see the “Superatom Limits”
section, on page 7-93.

Note:
This button is disabled if no Superatom Table has been loaded, see the
“Superatom Tables” section, on page 7-91.

Specifying Hot Symbols

Hot Symbols may be specified in the Parameters dialog, Symbol Parameters page Element
Limits frame, to allow their properties to be changed quickly and easily. Any element, isotope, or
superatom can be specified as a Hot Symbol as follows:

1. Select the box next to the required symbol.

2. Select the Symbol button; the Select Hot Symbol dialog is invoked.

Figure 7.73 The Select Hot Symbol dialog

3. Select the box for the required element, or isotope. The current element, or isotope, is
displayed in the Current Symbol frame.

Note:
By default, the list is sorted according to atomic number of the symbol. This can be changed
to alphabetical order by clicking on the Symbol column header. Clicking on the Atomic No.
column sorts back by atomic number again. Note that in alphabetical order, isotopes are still
sorted numerically.

4. Select the OK button; the Select Hot Symbol dialog is closed. The name on the Parameters
dialog, Symbol Parameters page, Symbol button will change to that selected in the Select
Hot Symbol dialog.

Page 7-88
 Spectrum

To Select a Hot Symbol

1. Check the box next to the required symbol.

2. If required, enter new values in the From and To boxes for the relevant symbol.

To Select an Element

Some common elements appear by default in the Parameters dialog, Symbol Parameters page;
to select one of these, select the relevant box.

To select an element that does not appear on the dialog:

1. Select the Periodic Table button; the Periodic Table dialog is invoked.

Note:
Selected elements are displayed in blue on the Periodic Table.

Figure 7.74 The Periodic Table dialog

2. Click on the required element to invoke the Elements & Isotopes dialog (see Figure 7.75);
this contains a list of isotopes for the element.

3. Check the first box in the list and select OK on each dialog until the display returns to the
Elemental Composition Window.

Note:
An element and its isotopes cannot be selected simultaneously; selecting the element will
deselect the isotopes.

To Select an Isotope

1. Select the Periodic Table button; the Periodic Table is invoked.

Note:
Elements with selected isotopes are displayed in blue on the Periodic Table.

Page 7-89
Spectrum

2. Select the required element; the Elements & Isotopes dialog is invoked. This contains a list
of isotopes for the element.

Figure 7.75 The Isotopes dialog

3. Select the boxes for the isotopes required, or select the Select Isotopes button to select all the
isotopes.

4. Select the OK button on each dialog until the display returns to the Elemental Composition
Window.

Note:
An element and its isotopes cannot be selected simultaneously, selecting the element will
deselect the isotopes.

To Deselect an Element

Some common elements appear in the Parameters dialog, Symbol Parameters page; to deselect
one of these deselect the relevant box, or select the Deselect All button to deselect all the elements.

To deselect an element that does not appear in the dialog:

1. Select the Periodic Table button; the Periodic Table dialog is invoked.

2. Select the required element; the Elements & Isotopes dialog is invoked.

3. Deselect the first box in the list.

4. Select the OK button on each dialog until the display returns to the Elemental Composition
Window.

To Deselect an Isotope

1. Select the Parameters dialog, Symbol Parameters page, Periodic Table button; the
Periodic Table dialog is invoked.

2. Select the required element; the Elements & Isotopes dialog is invoked.

Page 7-90
 Spectrum

3. Deselect the boxes for the isotopes that are not required.

4. Select the OK button on each dialog until the display returns to the Elemental Composition
Window.

To Change the Minimum and Maximum Values

For the elements displayed in the Parameters dialog, Symbol Parameters page, enter new values
in the From and To boxes for the relevant element. For other elements and for isotopes not
displayed in the Parameters dialog:

1. Select the Periodic Table button; the Periodic Table dialog is invoked.

2. Select the required element; the Elements & Isotopes dialog is invoked.

3. Click anywhere on the row, then on the Minimum or Maximum value, as required, and enter
a new value.

The Minimum value is the minimum number of elements or isotopes that the calculated
formula must contain; e.g. if the From value for Cl is 2 then the formula must contain Cl2, but
can contain any number above this, e.g. Cl3, Cl4, etc.

The Maximum value is the maximum number of elements or isotopes that the calculated
formula must contain; e.g. if the To value for Cl is 2 then the formula must contain Cl2, but
can contain any number below this.

If the Minimum and Maximum values are the same then the calculated formula must contain
this exact number of elements or isotopes; e.g. if the Minimum and Maximum values for Cl
are both 2, then the formula must contain Cl2.

4. Select the OK button on each dialog until the display returns to the Elemental Composition
Window.

Superatom Tables

General

A Superatom Table is an Access database containing details of large molecules that can be used in
the elemental composition search in the same way as elements and isotopes. The elements
database is loaded automatically on MassLynx start-up. The aminoacids.mdb database is also
supplied with MassLynx; it contains details of twenty common amino acids.

To Load a Superatom Database

1. Select the Parameters dialog, Symbol Parameters page, Superatoms frame, Tables button;
the Superatom Tables dialog is invoked. When this is first invoked, no database is loaded
and the list box will be empty.

2. Select the Add Superatom Table button; the Load Superatom Database browser is
invoked.

Page 7-91
Spectrum

Figure 7.76 The Superatom Tables dialog

3. Select the required database file.

4. Select the OK button; the Load Superatom Database browser is closed and the database file
is added to the Superatom Tables dialog.

To Create a New Superatom Database

Microsoft Access can be used to create new Superatom database tables, using the aminoacids.mdb
file as a template.

Note:
The table name, column headings and data types must match those in aminoacids.mdb.

Page 7-92
 Spectrum

Superatom Limits

General

Select the Parameters dialog, Symbol Parameters page, Superatoms frame, Limits button to
invoke the Superatom Limits dialog.

Figure 7.77 The Superatom Limits dialog

All superatoms are automatically switched on by default when a database is loaded. They are all
listed together in the order that the databases were loaded. If there is no more room left to
accommodate some or all of the symbols, they are not switched on. A warning is displayed telling
the User exactly which symbols could be enabled and how many currently enabled symbols will
have to be switched off in order to turn on the superatoms. A similar message will also be
displayed if an attempt is made to switch on elements or isotopes when the maximum number of
displayable symbols has been reached.

Figure 7.78 The Elemental Composition warning box

Page 7-93
Spectrum

To Select a Superatom

1. In the Superatom Limits dialog, select the boxes for the superatoms required, or select the
Select All button to select all superatoms.

2. Select the OK button on each dialog until the display returns to the Elemental Composition
Window.

To Deselect a Superatom

1. In the Superatom Limits dialog, deselect the boxes for the superatoms not required, or select
the Deselect All button to deselect all superatoms.

2. Select the OK button on each dialog until the display returns to the Elemental Composition
Window.

To Change the Minimum and Maximum Values

1. In the Superatom Limits dialog, double-click on the Minimum, or Maximum, value, as

required, and enter a new value.

The Minimum value is the minimum number of elements or isotopes that the calculated
formula must contain; e.g. if the From value for Cl is 2, the formula must contain Cl2, but can
contain any number above this, e.g. Cl3, Cl4, etc.

The value is the maximum number of elements or isotopes that the calculated formula must
contain; e.g. if the To value for Cl is 2 then the formula must contain Cl2, but can contain any
number below this.

If the Minimum and Maximum values are the same then the calculated formula must contain
this exact number of elements or isotopes; e.g. if the Minimum and Maximum values for Cl
are both 2, then the formula must contain Cl2.

2. Select the OK button on each dialog until the display returns to the Elemental Composition
Window.

Performing a Calibration
General

Calibration can be performed from the Spectrum Window.

To Make a New Calibration

1. Select the Spectrum Menu Bar Tools, Make Calibration command; the Make new
calibration dialog is invoked.

Page 7-94
 Spectrum

Figure 7.79 The Make new calibration dialog

2. Select a Reference file from the dropdown list box.

3. Select the Air references option to include air peaks at 28 and 32 in the calibration.

4. Select the Mass Measure button; the Mass Measure dialog is invoked. Enter the required
parameters. For more information see the “The Mass Measure Process” section, on
page 7-46.

5. Select OK. When processing is complete, the Calibration Report Window will be displayed.

Figure 7.80 The Calibration Report window

6. Parameters can be changed by selecting the Menu Bar Edit, Calibration Parameters
command; this invokes the Calibration Parameters dialog.

Page 7-95
Spectrum

Figure 7.81 The Calibration Parameters dialog

7. Change the required parameters and select the OK button to display the updated Calibration
Report.

8. Select the Calibration Report Window OK button to accept the calibration. A dialog
informing of a successful calibration is displayed.

9. Select the OK button.

To Apply a Calibration

1. Select the Spectrum Menu Bar Tools, Apply Calibration command; the Apply Calibration
dialog is invoked.

Figure 7.82 The Apply Calibration dialog

2. Select the OK button to apply the calibration. A dialog informing of a successful

recalibration is displayed.

3. Select the OK button.

Page 7-96
 Spectrum

To Modify a Calibration

1. Select the Spectrum Menu Bar Tools, Modify Calibration command. The Modify
Calibration dialog is invoked.

Figure 7.83 The Modify Calibration dialog

2. Change the Gain and/or Offset.

3. Select the OK button to apply the modification.

Lock Mass

The Lock Mass feature allows the User to specify a mass that will be located in the spectrum and
used to calculate an offset that can be applied to the rest of the spectrum. The Lock Mass dialog is
invoked by the Menu Bar Tools, Lock Mass command.

Figure 7.84 The Lock Mass dialog

Lock Mass The mass nearest to that specified in this text box [i.e. within the limits
(Da/e) specified in Window (Da/e)] will be located in the spectrum and used to
calculate an offset that can be applied to the rest of the spectrum.

Window (Da/e) Specifies the degree of error allowed when locating the Lock Mass.

Use When selected, the lock mass is matched to the nearest monoisotopic peak
Monoisotopic in the spectrum.
Peak (singly-
charged only)

Copying to and from the Windows Clipboard

General

The Windows Clipboard can be used to move data into, or out of, the Spectrum window, either as
a picture, or as a text list. For example, spectra or chromatograms can be pasted into reports
written with a Windows compatible word processor.

Page 7-97
Spectrum

The Spectrum Menu Bar Edit, Copy Picture command copies both a metafile and a bitmap to the
Clipboard. When the metafile is pasted into another Windows application, it can be re-scaled, if
required, without distorting the original image, as long as the original aspect ratio is maintained.

To Copy a Spectrum as a Picture to the Clipboard

1. Produce the required display in a Spectrum window.

2. Select the Tool Bar button, or select the Spectrum Menu Bar Edit, Copy Picture
command. The contents of the window are copied to the Clipboard as both a metafile and a
bitmap.

3. To read the image into another application as a metafile, select the application’s Edit, Paste
command. Alternatively, select the application’s Edit, Paste Special command to have the
option of pasting either the metafile or the bitmap image.

To Copy a Spectrum as a Text List to the Clipboard

1. Display the required mass range in a Spectrum window.

2. Select the Tool Bar button, or select the Spectrum Menu Bar Edit, Copy Spectrum List
command. The displayed section of the spectrum will be copied to the Clipboard as (mass,
intensity) pairs.

3. To read the information into another application, select the application’s Edit, Paste
command.

To Paste Information from the Windows Clipboard into a

Spectrum Window

1. Select the Tool Bar button, or select the Spectrum Menu Bar Edit, Paste command to
paste the default Clipboard object to Spectrum. Choose the Edit, Paste Special command to
choose which object to paste into the Spectrum. These objects would typically be metafiles,
bitmaps, or text.

2. Use the mouse to drag the outline of the image to the required position.

Any contents of the Clipboard, be it a bitmap, a metafile or text, can be pasted into a Spectrum
window. If the data is in textual or metafile form, it can be re-scaled using the mouse, and there
will be no distortion of the image. However, if a bitmap is pasted, re-scaling is done by stretching
the image; this will cause some distortion. To avoid this, scale the image to the required size
before copying it to the Clipboard.

Removing Pasted Input from the Display

1. Click on item to be removed.

2. Press the keyboard Delete key.

Page 7-98
 Spectrum

Manipulating Library Spectra

To Display a Library Entry

1. Select the Spectrum Menu Bar Edit, Library, Get Spectrum command. The Display
Library Spectrum dialog is invoked.

Figure 7.85 The Display Library Spectrum dialog

2. If required, select a new library by selecting the File button. The Open Browser is invoked;
select the required library file and select the OK button.

3. Specify an entry number in the Display Library Spectrum dialog Entry: text box.

4. The library spectrum may be added to the current spectrum window, replace the current
spectrum, or be placed in a new window. Select the Add trace, Replace trace or New
window option as appropriate.

5. Select the OK button.

Once a spectrum from a library has been displayed, the rest of the library may be browsed by using
the Spectrum Tool Bar button.

To Append the Current Spectrum to the Current Library

1. Select the Spectrum Menu Bar Edit, Library, Append command. The Append Spectrum
dialog is invoked.

Figure 7.86 The Append Spectrum dialog

2. If required, select a new library by selecting the File button. The Append File Select
Browser is invoked; select the required library file and select the OK button.

3. Select the OK button.

Page 7-99
Spectrum

Page 7-100
 Strip and Combine Functions

Chapter 8 Strip and Combine

Functions

Page 8-1
Strip and Combine Functions

Contents
The Strip Program ......................................................................................................................... 8-5
General............................................................................................................................... 8-5
The Strip Datafile Dialog ................................................................................................... 8-6
The Strip Datafile Dialog Menu Bar .................................................................................. 8-7
Creating an Enhanced Data File......................................................................................... 8-8
Creating a Subtracted Data File ....................................................................................... 8-10
Creating a Clustered Data File ......................................................................................... 8-11
Creating a CODA Data File ............................................................................................. 8-12
Selecting an Input Data File to Process............................................................................ 8-13
Selecting a Sub-range to Process ..................................................................................... 8-14
Selecting a Background Data File .................................................................................... 8-15
Selecting an Output Data File .......................................................................................... 8-16
Setting Subtract Datafile Options..................................................................................... 8-16
Setting Enhance Datafile Options .................................................................................... 8-17
Setting Cluster Datafile Options ...................................................................................... 8-18
Setting Cluster Centroid Options ..................................................................................... 8-19
Setting CODA Options .................................................................................................... 8-21
Stopping a Process ........................................................................................................... 8-21
The Combine Functions Program................................................................................................ 8-21
General............................................................................................................................. 8-21
Creating a Combined Data File........................................................................................ 8-22
The Combine Datafile Functions Dialog Menu Bar......................................................... 8-23
Selecting an Input Data File ............................................................................................. 8-24
Selecting a Sub-range to Process ..................................................................................... 8-25
Selecting an Output Data File .......................................................................................... 8-26
Stopping a Process ........................................................................................................... 8-26
The Combine All Files Program.................................................................................................. 8-26
General............................................................................................................................. 8-26
The Combine All Files Dialog Input File(s) List ............................................................. 8-28
The Combine All Files Dialog Menu Bar ........................................................................ 8-29
Processing Files................................................................................................................ 8-29

Illustrations
Figure 8.1 The Strip Datafile dialog............................................................................................. 8-6
Figure 8.2 The Strip Datafile Dialog File Menu........................................................................... 8-7
Figure 8.3 The Options Menu ...................................................................................................... 8-7
Figure 8.4 The Strip Datafile dialog Help menu .......................................................................... 8-8
Figure 8.5 Chromatogram showing Ribonuclease tryptic digest raw data file
(lower trace) and enhanced data file (upper trace)...................................................... 8-8
Figure 8.6 A single spectrum from a Ribonuclease tryptic digest raw data file
(lower trace) and enhanced data file (upper trace)...................................................... 8-9
Figure 8.7 Chromatogram showing Fentuin tryptic digest raw data file (lower
trace) and enhanced data file (upper trace) ................................................................. 8-9
Figure 8.8 Chromatogram showing V50 raw data file (lower trace) and subtracted
data file (upper trace)................................................................................................ 8-10
Figure 8.9 Chromatogram showing blank data file (bottom trace), sample data
file (middle trace) and subtracted data file (top trace) .............................................. 8-11
Figure 8.10 Spectrum showing mixture of two chlorines data file (lower trace)
and cluster analysed data file to show mass difference of 2 amu (upper trace) ...... 8-12
Figure 8.11 Chromatogram showing raw TIC data (lower trace) and CODA data
(upper trace). The CODA data exhibits none of the spikes seen in the raw
trace, and lower background................................................................................... 8-13
Figure 8.12 The Input Datafile dialog ........................................................................................ 8-13

Page 8-2
 Strip and Combine Functions

Figure 8.13 The Strip Data Browser dialog ............................................................................... 8-14

Figure 8.14 The Subtract Background File dialog ..................................................................... 8-15
Figure 8.15 Output File dialog ................................................................................................... 8-16
Figure 8.16 The Subtract Datafile Options dialog...................................................................... 8-16
Figure 8.17 The Enhance Datafile Options dialog ..................................................................... 8-17
Figure 8.18 The Cluster Analysis Options dialog ...................................................................... 8-18
Figure 8.19 The Fast Centroid dialog......................................................................................... 8-20
Figure 8.20 The CODA dialog................................................................................................... 8-21
Figure 8.21 Spectra from a protease digest of Histone............................................................... 8-22
Figure 8.22 The Combine Datafile Functions dialog ................................................................. 8-23
Figure 8.23 The Combine Datafile Functions Dialog File Menu............................................... 8-23
Figure 8.24 The Combine Datafile Functions Dialog Help Menu ............................................. 8-24
Figure 8.25 The Input Datafile dialog ........................................................................................ 8-24
Figure 8.26 The Strip Data Browser dialog ............................................................................... 8-25
Figure 8.27 Output File dialog ................................................................................................... 8-26
Figure 8.28 The Combine All Files dialog................................................................................. 8-27
Figure 8.29 The Combine All Files Dialog File menu ............................................................... 8-29
Figure 8.30 The Combine All Files Dialog Operations menu.................................................... 8-29

Page 8-3
Strip and Combine Functions

Page 8-4
 Strip and Combine Functions

The Strip Program

General

The Strip program provides a way of removing unwanted background and noise from a data file.
Processing a data file using Strip creates a new file which is stored in the same format as a raw
data file and can be displayed and processed in the same way as a raw data file. The original input
file is retained unmodified.

Strip provides four processing options:

• Enhance

Removes noise from continuum data files. It examines each data point, and its close
neighbors, to determine whether it is noise, or part of a real feature. Data points not
considered to be valid are removed from the output data file. Enhance can significantly
reduce data file size.

• Subtract

This option can subtract either a single background spectrum, or a whole data file from the
input file. Processed spectra can be subtracted, enabling averaged spectra to be used as
background. Both centroid and continuum type files can be subtracted, however, different
types cannot be mixed.

• Cluster

Detects pairs, or triplets, of peaks separated by a specified mass difference. Parameters

specified are mass differences and expected intensity ratios (both with tolerances), together
with a time window and a global threshold. The resulting data file will contain only these
peaks. Again, Cluster will significantly reduce data file size.

• CODA (COmponent Detection Algorithm)

Essentially removes mass chromatograms that represent background noise from the dataset.
Each raw mass chromatogram is compared to a smoothed, standardized mass chromatogram,
and masses in which the background noise is high, or in which spikes are present, are rejected.

The Strip program is accessed via the Strip Datafile dialog, see the “The Strip Datafile Dialog”
section, on page 8-6.

Note:
The Strip program cannot be accessed while the Combine Functions program is loaded. Likewise,
the Combine Functions program cannot be accessed while the Strip program is loaded.

Page 8-5
Strip and Combine Functions

The Strip Datafile Dialog

The Strip Datafile dialog is invoked by selecting the MassLynx Tools Shortcut Bar Strip icon. A
Menu Bar is provided at the top of the dialog, refer to the “The Strip Datafile Dialog Menu Bar”
section, on page 8-7 for details.

Figure 8.1 The Strip Datafile dialog

Input Frame The data file name and function number currently selected for processing
are displayed in this frame.

Input button Invokes the Input Datafile dialog, see the “Selecting an Input Data File to
Process” section, on page 8-13.

Background The data file, function and scan number to be used as background when
Frame performing the Subtract process are displayed in this frame.

Background Invokes the Subtract Background File dialog, see the “Selecting a
button Background Data File” section, on page 8-15.

Output Frame The name of the data file that will be created when processing has been
completed is displayed in this frame.

Output button Invokes the Output File dialog, see the “Selecting an Output Data File”
section, on page 8-16.

Process button Starts the selected process.

Stop button Stops the current process.

Page 8-6
 Strip and Combine Functions

Processes Frame Used to select the processing option.

Enhance Selects the Enhance process.

Subtract Selects the Subtract process.

Cluster Selects the Cluster process.

CODA Selects the CODA process.

(COmponent
Detection
Algorithm)

The Strip Datafile Dialog Menu Bar

The File Menu

Figure 8.2 The Strip Datafile Dialog File Menu

Input Invokes the Strip Data Browser dialog, see the “Selecting an Input Data
File to Process” section, on page 8-13.

Background Invokes the Subtract Background File dialog, see the “Selecting a
Background Data File” section, on page 8-15.

Exit Closes the application.

The Options Menu

Figure 8.3 The Options Menu

Enhance datafile Invokes the Enhance Datafile Options dialog, see the “Setting Enhance
options Datafile Options” section, on page 8-17.

Subtract datafile Invokes the Subtract Datafile Options dialog, see the “Setting Subtract
options Datafile Options” section, on page 8-16.

Cluster datafile Invokes the Cluster Analysis Options dialog, see the “Setting Cluster
options Datafile Options” section, on page 8-18.

CODA options Invokes the CODA dialog, see the “Setting CODA Options” section, on
page 8-21.

Page 8-7
Strip and Combine Functions

The Help Menu

Figure 8.4 The Strip Datafile dialog Help menu

Help on Strip Invokes the help utility for the Strip application.
processing

Creating an Enhanced Data File

1. Select the Strip Datafile dialog Enhance option.

2. Select the required Input file and the sub-range to process. See the “Selecting an Input Data
File to Process” section, on page 8-13.

3. Select the required Output file. See the “Selecting an Output Data File” section, on page 8-16.

4. Select the Menu Bar Options, Enhance data file Options command to set the Enhance
options. See the “Setting Enhance Datafile Options” section, on page 8-17.

5. Select the Process button to start processing the data file. The status bar at the bottom of the
Strip dialog displays the progress of the current process.

Figure 8.5 shows two chromatogram traces. The lower trace is a raw data file obtained from a
Ribonuclease tryptic digest. The upper trace shows the same data file after it has been processed
using the Enhance option. The background noise level has been greatly reduced in the enhanced
data. The original data file size of 19 MB has been reduced to 0.5 MB in the enhanced data file.

Ribonuclease tryptic digest.

TH01R Scan ES+
100 31.56 TIC
1.41e9
Area

29.01
%
20.17 25.87
1.64 12.18 16.5217.37
21.11
8.87 35.73 39.47 44.91
0
TH01 Scan ES+
100 31.14 TIC
1.89e9
Area

1.64 29.01
%
20.09
16.5217.37 21.11 25.87
12.18
15.92
8.95 11.59 35.73 39.47 44.91
0 rt
5 00 10 00 15 00 20 00 25 00 30 00 35 00 40 00 45 00 50 00
Figure 8.5 Chromatogram showing Ribonuclease tryptic digest raw data file (lower trace)
and enhanced data file (upper trace)

Page 8-8
 Strip and Combine Functions

Figure 8.6 shows part of a single scan from the raw and enhanced data files. The background
noise in the enhanced spectrum has been greatly reduced.

Ribonuclease tryptic digest.

TH01R 384 (32.667) Scan ES+
100 1.76e7
1711

1521
1369 1712
1521
%
1710

1371 1491 1504 1525 1709 1713

1647 1656
0
TH01 384 (32.667) Scan ES+
100 1.76e7
1711

1521
1369
1521 1712
%
1710

13711390 1491 1504 1525 1547 1709 1713

1647 1656 1737 1769
0 Da/e
1350 1400 1450 1500 1550 1600 1650 1700 1750
Figure 8.6 A single spectrum from a Ribonuclease tryptic digest raw data file (lower trace)
and enhanced data file (upper trace)

Figure 8.7 shows the chromatogram of a Fentuin tryptic digest. The lower trace is the raw data
file, the upper trace shows the same data file after it has been processed using the Enhance option
to reduce background noise. The original data file size of 41.2 MB has been reduced to 1.4 MB in
the enhanced data file.

Fetuin tryptic digest, 200pmole,

FET01C Scan ES+
32.43 TIC
100
1.23e8
20.55 Area

42.55
7.59
%
9.34 13.26 26.22 30.54 35.80 51.33
6.10 18.39 58.35
24.33 39.18 47.55 54.84
3.54
0
FET01 Scan ES+
32.43 TIC
100
2.00e8
20.55 Area
42.55
7.59
26.22 30.54 35.80 51.33
9.34 13.26 42.15 43.77 47.41 54.84 58.35
6.10 18.39 24.33
%
3.54 60.37 61.32

0 Scan
25 50 75 100 125 150 175 200 225 250 275 300 325 350 375 400 425 450
Figure 8.7 Chromatogram showing Fentuin tryptic digest raw data file (lower trace) and
enhanced data file (upper trace)

Page 8-9
Strip and Combine Functions

Creating a Subtracted Data File

1. Select the Strip Datafile dialog Subtract option.

2. Select the required Input file and the sub-range to process; see the “Selecting an Input Data
File to Process” section, on page 8-13.

3. Select the required Background file; see the “Selecting a Background Data File” section, on
page 8-15.

4. Select the required Output file; see the “Selecting an Output Data File” section, on page 8-16.

5. Set the Subtract options by selecting the Menu Bar Options, Subtract datafile options
command; see the “Setting Subtract Datafile Options” section, on page 8-16.

6. Select the Process button to start processing the data file. The status bar at the bottom of the
Strip Datafile dialog displays the progress of the current process.

The lower trace in Figure 8.8 shows the TIC chromatogram of the V50 data file. The upper trace
shows the TIC chromatogram of the same data file after a background scan (scan 761 at retention
time 32 minutes) has been subtracted.

50ppm. Volatile Organic Analysis Calibration Standard.

V50A Scan EI-
100 0.99 TIC
6.53e5
Area

19.61
26.20 27.58 30.38
7.67 14.10 15.98 21.99 24.87
10.43 13.35 34.0536.06
37.85
2.03
0
V50 Scan EI-
0.99 TIC
100 6.69e5
Area

%
19.61 27.58
24.87 26.20 30.38
7.67 14.10 15.98 21.99 37.85
10.43 13.35 34.0536.06
2.03 3.20
0 rt
5.00 10.00 15.00 20.00 25.00 30.00 35.00
Figure 8.8 Chromatogram showing V50 raw data file (lower trace) and subtracted data file
(upper trace)

Page 8-10
 Strip and Combine Functions

Figure 8.9 shows an example of subtracting a complete data file from another data file. The
bottom trace shows a mass chromatogram from the “blank” sample, the middle trace shows a mass
chromatogram from the analyte sample and the top trace shows the result of subtracting the
“blank” data file from the analyte data file.

LMD25A SIR of 2 Channels ES+

100 706.30
10.61 3.77e5
Area

%
8.03
14.32
1.58 14.76
0
LMD25 SIR of 2 Channels ES+
100 10.61 706.30
3.77e5
Area

% 8.03 14.32
15.94
1.60 4.17 17.66
0
LMD12 SIR of 2 Channels ES+
100 706.30
3.77e5
Area

% 15.79 17.35 18.96

6.78 14.06
11.27
1.71 4.19
0 rt
2 00 4 00 6 00 8 00 10 00 12 00 14 00 16 00 18 00 20 00 22 00
Figure 8.9 Chromatogram showing blank data file (bottom trace), sample data file (middle
trace) and subtracted data file (top trace)

Creating a Clustered Data File

1. Select the Strip Datafile dialog Cluster option.

2. Select the required Input file and the sub-range to process, see the “Selecting an Input Data
File to Process” section, on page 8-13.

3. Select the required Output file, see the “Selecting an Output Data File” section, on page 8-16.

4. Select the Menu Bar Options, Cluster data file Options command to set the Cluster options.
See the “Setting Cluster Datafile Options” section, on page 8-18.

5. Select the Process button to start processing the data file. The status bar at the bottom of the
Strip dialog displays the progress of the current process.

Figure 8.10 shows the spectrum of a mixture of two chlorines. The lower trace is the raw data file,
the upper trace shows the same data file after it has been processed using the Cluster option to
show pairs of peaks differing in mass by 2 Da. The original data file size of 110 Kb has been
reduced to 152 bytes (peaks only) in the clustered data file.

Page 8-11
Strip and Combine Functions

Scan ES-
183.0078 1.06e5
100

%
155.0078

157.0313 185.0391
0
Scan ES-
183.0078 1.06e5
100

%
155.0078

191.0234
209.0234
245.0547 263.0469
281.0859 335.1484
317.1016 353.1250 389.0859 407.1172
425.1250 460.9688
0 Da/e
160 180 200 220 240 260 280 300 320 340 360 380 400 420 440 460 480 500

Figure 8.10 Spectrum showing mixture of two chlorines data file (lower trace) and cluster
analysed data file to show mass difference of 2 amu (upper trace)

Creating a CODA Data File

1. Select the Strip Datafile dialog CODA option.

2. Select the required Input file to process. See the “Selecting an Input Data File to Process”
section, on page 8-13.

Note:
The input mass range can’t be changed and all functions are processed irrespective of
function selected.

3. Select the required Output file. See the “Selecting an Output Data File” section, on page 8-16.

4. Select the Menu Bar Options, CODA Options command to set the CODA options. See the
“Setting CODA Options” section, on page 8-21.

5. Select the Process button to start processing the data file. The status bar at the bottom of the
Strip dialog displays the progress of the current process.

Page 8-12
 Strip and Combine Functions

6.0 femtomoles fetuin tryptic lcms 50 um i.d column with 5/2 etoh/prop/formic solvents
UCSF20C 1: Scan ES+
TIC
100
6.48e8

30.40

%
38.06
44.04

44.42
52.38 53.44 56.54 60.86 64.57 67.53 69.12
45.10
0
UCSF20 1: Scan ES+
52.45 67.61 TIC
100 44.04
6.48e8
48.82 53.51
30.48
45.56
53.67
56.62 63.43 66.62
49.12
38.13
49.58 69.65
%
37.15 38.89
21.61
41.47
29.57 33.82
16.00 19.42 24.64
0.17 8.35 11.54
14.42

0 Scan
50 100 150 200 250 300 350 400 450 500 550 600 650 700 750 800 850 900

Figure 8.11 Chromatogram showing raw TIC data (lower trace) and CODA data (upper
trace). The CODA data exhibits none of the spikes seen in the raw trace, and lower
background.

Selecting an Input Data File to Process

The data file name and function number currently selected for processing are displayed in the
Strip Datafile dialog Input frame.

To change the current input file:

1. Either:

a. Select the Strip Datafile dialog Input button, the Input Datafile dialog is invoked.

Figure 8.12 The Input Datafile dialog

b. Select the File button, the Strip Data Browser dialog is invoked.

Or:

Select the Strip Datafile dialog Menu Bar File, Input command, the Strip Data
Browser dialog is invoked.

Page 8-13
Strip and Combine Functions

Figure 8.13 The Strip Data Browser dialog

2. Select the required file.

3. Select the required Function from the drop down list box.

4. Select OK to return to the Input Datafile dialog.

5. By default the whole of the selected function will be processed; to specify sub-ranges see the
“Selecting a Sub-range to Process” section, below.

6. To strip all the functions for a data file, select the Input Datafile dialog Strip All option.

7. When an input file is selected a default output file name is displayed on the Strip Datafile
dialog Output frame. This can be changed, see the “Selecting an Output Data File” section,
on page 8-16, for details.

Selecting a Sub-range to Process

Note:
CODA does not allow sub-range selection. This is because all functions in the dataset are
processed.

Processing a mass, or retention time, sub-range of the input file has the advantages of reducing
both processing time and the size of the resulting output file.

Page 8-14
 Strip and Combine Functions

In the Input Datafile dialog, enter values for the Mass Range and Retention Time range that are
to be processed. These ranges can also be set from Spectrum and Chromatogram by using the
mouse to identify the desired range.

To set the Mass Range parameters using the mouse, press the right mouse button at one end of the
Spectrum region of interest and, without releasing the button, drag the mouse horizontally to the
other end. A “rubber band” is stretched out to indicate the selected range. The Input Datafile
dialog will be updated to show the new Mass Range.

To set the Retention Time parameters using the mouse, press the right mouse button at one end of
the Chromatogram region of interest and, without releasing the button, drag the mouse
horizontally to the other end. A “rubber band” is stretched out to indicate the selected range. The
Input Datafile dialog will be updated to show the new Retention Time range.

The Input Datafile dialog Default button sets both Mass Range and Retention Time range to the
full range of the current file.

Selecting a Background Data File

The data file, function and scan number to be used as background when performing the Subtract
process are displayed in the Strip Datafile dialog Background frame. Previously processed
spectra can be used as background.

Note:
The background file is not used for Enhanced processing.

1. Select the Strip Datafile dialog Background button; the Subtract Background File dialog is
invoked.

Figure 8.14 The Subtract Background File dialog

2. Select the File button; the Strip Data Browser dialog is invoked.

3. Select the required file.

4. Select the required Function from the drop down list box.

5. To select processed data, select the History button and highlight the required process.

Note:
Only saved processes can be selected.

6. Select OK to return to the Subtract Background File dialog.

Page 8-15
Strip and Combine Functions

7. If a single background scan is to be subtracted, enter the scan number in the Background
Scan text box. The scan number can also be set from Spectrum and Chromatogram by
right-clicking with the mouse to identify the desired scan.

8. If the whole of the background file is to be subtracted, select the Use all background file
check box. In this case the background scan with the closest retention time to each input scan
will be subtracted.

Note:
The Background Scan text box is disabled when the Use all background file option is selected.

Selecting an Output Data File

The name of the data file that will be created when processing has been completed is displayed in
the Strip Datafile dialog Output frame.

When an Input file is selected, the Output file defaults to the same directory and a name based
upon the Input file name, with an extra letter appended. For example, if the input file was
\masslynx\data\v50.raw the default output file might be \masslynx\data\v50a.raw. When
defaulting the output name, MassLynx attempts to choose a name that doesn’t already exist.

To change the default output file and directory select the Strip Datafile dialog Output button.
The Output File dialog is invoked.

Figure 8.15 Output File dialog

Enter a Name: for the output file. To change the file directory, enter the full path name of the file.

Setting Subtract Datafile Options

To set the Subtract processing parameters, select the Strip Datafile dialog Menu Bar Options,
Subtract datafile options command; the Subtract Datafile Options dialog is invoked.

Figure 8.16 The Subtract Datafile Options dialog

Page 8-16
 Strip and Combine Functions

Peak Width This parameter is the spectral peak width in amu; it is only used when
(amu, centroid subtracting centroid data. The peak width can be determined from
only) inspection of the tune peaks in the tune page. The peak width is used to
determine if peaks present in the input and background data represent the
same peak.

Background This is applied to the intensities of the peaks in the background spectra
multiplication before they are subtracted from peaks in the input spectra. This provides a
factor method of adjusting the height of the subtracted background.

Default Sets the parameters to their default values.

Setting Enhance Datafile Options

To set the Enhance processing parameters select the Strip Datafile dialog Menu Bar Options,
Enhance datafile options command; the Enhance Datafile Options dialog is invoked.

Figure 8.17 The Enhance Datafile Options dialog

Enhance operates on continuum data only, it works by examining each spectrum data sample to
determine if it is a noise spike or part of a real feature. This is achieved by examining neighboring
samples on the mass scale and at the same area in the preceding and following scans.

Window Frame

Mass (data Determines how many samples to examine each side of the current sample
points ±) along the mass scale. It should not exceed half the number of samples that
make up a peak.

Scans (±) Determines how many scans to examine each side of the current scan. It
should not exceed half the number of scans a chromatogram peak is
present.

Spike Rejection
Frame
Intensity Defines an absolute intensity that a data point must exceed to be regarded
Threshold as being significant. For spectra with a high baseline this parameter will
need adjusting so that its value is approximately equal to the intensity at
the top of the noise. The larger this value, the more likely that information
will be discarded as being noise.

Page 8-17
Strip and Combine Functions

Minimum Determines the minimum percentage of neighboring samples examined

Abundance whose intensity must be above the specified threshold for the current
(%) sample not to be rejected as noise. The larger this value the more likely
that a sample will be discarded.

Default Sets the parameters to their default values.

For example, if Mass (data points ±) is set to 2, two samples either side of the current sample will
be examined, including the current sample, making five in all. If Scans (±) is set to 1, one scan
either side of the current scan will be used, so (including the current scan), three scans will be
used. Multiplying the number of scans by the number of samples in each scan shows that fifteen
samples are examined in total. Consequently, for a sample to be accepted, 60% of these samples
(nine samples) must have an intensity greater than the specified Intensity Threshold.

Setting Cluster Datafile Options

To set the Cluster processing parameters, select the Strip Datafile dialog Menu Bar Options,
Cluster datafile options command; the Cluster Analysis Options dialog is invoked.

Figure 8.18 The Cluster Analysis Options dialog

Cluster operates on both centroid and continuum data. For continuum data, a special “fast
centroid” conversion process is used prior to the cluster processing, see the “Setting Cluster
Centroid Options” section, on page 8-19. Cluster works by examining each pair (or triplet) of
peaks in each spectrum to determine if they are separated by the correct mass difference(s), and if
their intensity ratios lie in the correct range(s). If the Time Window parameter is set to a value
other than zero, the neighboring scans within that time window (±) are examined.

Mass 1 Frame

First Mass Determines the requested separation and intensity ratio of the first pair of
Diff… amu peaks. The intensity ratio is calculated as (intensity of low mass
and First Ratio peak/intensity of high mass peak). The requested intensity ratio may be
less than one. Intensity ratio comparison can be disabled by deselecting
the Use Intensity Ratios option, see below.

Page 8-18
 Strip and Combine Functions

Mass 2 Frame

Use Second Disables the second mass difference. If this option is not selected,
Mass examination is restricted to pairs of peaks only, not triplets.
Difference
Second Mass Determines the mass difference between and intensity ratio of the first and
Diff… amu and third peaks in the triplet (not the first and second).
Second Ratio
Mass Tolerance Specifies a window (±) for each of the specified mass differences
amu (maximum of two). Pairs, or triplets, of peaks are detected if the
corresponding peak(s) lie at the specified mass difference ± the specified
mass tolerance.

Ratio Specifies the maximum mismatch between specified and calculated

Tolerance… % intensity ratios. It is specified as a percentage of the intensity ratio(s).

Time Window… Determines how far apart scans may lie in which peaks forming part of the
min pair/triplet are located. For instance, if the Time Window is ± 0.5 min,
with mass difference 5.0 amu, then a peak at mass 25.0 Da in a scan at
time 2.2 min will match with a peak at mass 30.0 Da in a scan at time
2.7 min.

Threshold Defines an absolute intensity that a data point must exceed to be regarded
as being significant. For spectra with a high baseline this parameter will
need adjusting so that its value is approximately equal to the intensity at
the top of the noise. The larger this value, the more likely that information
will be discarded as being noise.

Use Intensity Enables Intensity ratio comparison; if this option is not selected, no ratio
Ratios comparison is attempted, and peaks are selected purely on the grounds of
mass difference.

Default Sets the parameters to their default values.

Centroid Invokes the Fast Centroid dialog, see the “Setting Cluster Centroid
Options” section, below.

For example, using the values in the above dialog; cluster will detect triplets, with the mass
difference between the first two peaks being 21.5-22.5 Da, and between the first and third peaks
31.5-32.5 Da. The intensity ratios of the peaks must lie in the range 0.7 to 1.3 (low mass
peak/high mass peak), and the peaks must lie in the same scan. The peaks must be more intense
than 0.01% times the most intense peak in the function.

Setting Cluster Centroid Options

For continuum data, a special “fast centroid” conversion process is used prior to the cluster
processing. The Fast Centroid process is unique to the cluster algorithm. It reduces the time taken
to centroid each scan of a LC run. Consequently, it will not deal as accurately with multiplets as
the standard centroid algorithm (see Chapter 7, “Spectrum”), but should be perfectly adequate for
most applications. The calculation can be performed in three ways:

• Select the most intense (top) point on the peak. This method is the least prone to errors
caused by unresolved adducts in ElectroSpray spectra.

• Calculate the centroid of the peak. This is equivalent to finding the vertical line passing
through the center of gravity of the peak. This will provide a more accurate mass
measurement, unless the peak contains unresolved adducts.

Page 8-19
Strip and Combine Functions

• Calculate the median of peak area. This is equivalent to drawing the vertical line such that
half the area of the peak lies on either side.

To set the Cluster “fast centroid” conversion process parameters, select the Cluster Analysis
Options dialog Centroid button (see the “Setting Cluster Datafile Options” section, on
page 8-18); the Fast Centroid dialog is invoked.

Figure 8.19 The Fast Centroid dialog

Center method
Frame
Peak width at Specifies the expected width of the continuum peaks at baseline. It has
base (amu) two purposes

• It determines the amount of smoothing that is applied to the

continuum spectrum prior to centroiding proper.

• It determines how close together two bars must lie in order to be

“grouped” into a single bar, i.e. it controls the multiplet resolution.

For smoothing, the width at the half height of the peak is estimated as half
the specified width at baseline, and it is this estimated value that is used in
the smooth. For multiplet resolution, peaks closer together than the
specified Peak width at base distance will be regarded as a singlet.

Baseline Specifies the minimum signal level in the spectrum above which a peak
threshold will be considered significant.

Top Selects the “top” calculation method.

Centroid top Selects the “centroid” calculation method.

(%)
Median Selects the “median” calculation method.

Centered
spectrum Frame

Page 8-20
 Strip and Combine Functions

Areas When selected, the height of the bars represents the sum of the intensities
of the points across the peak in the continuum trace.

Heights When selected, the height of the bars represents the intensity of the
continuum trace at the mass of the bar.

Setting CODA Options

CODA operates on both centroid and continuum data. It works by standardizing and smoothing
each mass chromatogram in the dataset, then comparing the smoothed, standardized, mass
chromatogram with the raw chromatogram. If they are sufficiently similar, as determined by the
MCQ Index parameter, the mass chromatogram is preserved, otherwise it is removed.
Essentially, mass chromatograms that contain spikes, or are noisy, will be dissimilar after
smoothing and standardization to the raw mass chromatogram, and are hence rejected.

To set the CODA processing parameters, select the Strip Datafile dialog Menu Bar Options,
CODA options command; the CODA dialog is invoked.

Figure 8.20 The CODA dialog

MCQ Index Specifies how similar the smoothed, standardized mass chromatogram
must be to the raw mass chromatogram before it is preserved. The
parameter is in the range 0 to 1 inclusive; a value of 0 will preserve all
mass chromatograms and result in the raw file being copied to the output.
A value of 1 will result in all mass chromatograms being rejected, and an
empty file. Values around the default value of 0.75 are most useful, with
the range 0.65 to 0.85 recommended.

Smooth Specifies the amount of smoothing given to raw mass chromatograms.

window… points The default value of 5 is usually adequate. This window is ± a number of
data points around the central point.

Stopping a Process

To stop a process before it has reached completion, select the Strip Datafile dialog Stop button.
Confirmation of the action will be requested.

The output data file will contain all the information written up to the point at which the process
was stopped.

The Combine Functions Program

General

The Combine Functions program combines all functions in a data file to produce a new data file
containing a single function, which is the sum of the multiple functions. To use the Combine
Functions option all the functions in the data file must have been acquired using the same scan

Page 8-21
Strip and Combine Functions

range and scan rate, or must contain the same SIR or MRM channels. The Combine Functions
option is particularly useful for combining functions acquired with different values of cone voltage
or collision energy

The figure below shows data from a protease digest of Histone. The lower traces show functions
acquired at different values of collision energy. The data file was processed using the Combine
Functions option to give the combined data file shown in the upper trace.

Figure 8.21 Spectra from a protease digest of Histone

Creating a Combined Data File

Select the MassLynx Tools Shortcut Bar Combine Functions icon; this invokes the Combine
Datafile Functions dialog. A Menu Bar is provided at the top of the dialog, refer to the “The
Combine Datafile Functions Dialog Menu Bar” section, on page 8-23, for details.

Note:
The Combine Functions program cannot be accessed while the Strip program is loaded.
Likewise, the Strip program cannot be accessed while the Combine Functions program is
loaded.

Page 8-22
 Strip and Combine Functions

Figure 8.22 The Combine Datafile Functions dialog

Input Frame The data file name and function number currently selected for processing
are displayed in this frame.

Input button Invokes the Input Datafile dialog, see the “Selecting an Input Data File”
section, on page 8-24.

Output Frame The name of the data file that will be created when processing has been
completed is displayed in this frame.

Output button Invokes the Output File dialog, see the “Selecting an Output Data File”
section, on page 8-26.

Process button Starts the selected process.

Stop button Stops the current process.

The Combine Datafile Functions Dialog Menu Bar

The File Menu

Figure 8.23 The Combine Datafile Functions Dialog File Menu

Input Invokes the Strip Data Browser dialog, see the “Selecting an Input Data
File” section, on page 8-24.

Exit Closes the application.

Page 8-23
Strip and Combine Functions

The Help Menu

Figure 8.24 The Combine Datafile Functions Dialog Help Menu

Help on Invokes the help utility for the Combine Functions application.
Combine
Functions

Selecting an Input Data File

The data file name and function number currently selected for processing are displayed in the
Combine Datafile Functions dialog Input frame.

To change the current input file:

1. Either:

a. Select the Combine Datafile Functions dialog Input button, the Input Datafile dialog is
invoked.

Figure 8.25 The Input Datafile dialog

b. Select the File button, the Strip Data Browser dialog is invoked.

Or:

Select the Combine Datafile Functions dialog Menu Bar File, Input command, the Strip
Data Browser dialog is invoked.

Page 8-24
 Strip and Combine Functions

Figure 8.26 The Strip Data Browser dialog

2. Select the required file.

3. Select the required Function from the drop down list box.

4. Select OK to return to the Input Datafile dialog.

5. By default the whole of the selected function will be processed; to specify sub-ranges see the
“Selecting a Sub-range to Process” section, below.

6. When an input file is selected a default output file name is displayed on the Strip Datafile
dialog Output frame. This can be changed, see the “Selecting an Output Data File” section,
on page 8-26, for details.

Selecting a Sub-range to Process

Processing a mass, or retention time, sub-range of the input file has the advantages of reducing
both processing time and the size of the resulting output file.

In the Input Datafile dialog, enter values for the Mass Range and Retention Time range that are
to be processed. These ranges can also be set from Spectrum and Chromatogram by using the
mouse to identify the desired range.

To set the Mass Range parameters using the mouse, press the right mouse button at one end of the
Spectrum region of interest and, without releasing the button, drag the mouse horizontally to the

Page 8-25
Strip and Combine Functions

other end. A “rubber band” is stretched out to indicate the selected range. The Input Datafile
dialog will be updated to show the new Mass Range.

To set the Retention Time parameters using the mouse, press the right mouse button at one end of
the Chromatogram region of interest and, without releasing the button, drag the mouse
horizontally to the other end. A “rubber band” is stretched out to indicate the selected range. The
Input Datafile dialog will be updated to show the new Retention Time range.

The Input Datafile dialog Default button sets both Mass Range and Retention Time range to the
full range of the current file.

Selecting an Output Data File

The name of the data file that will be created when processing has been completed is displayed in
the Combine Datafile Functions dialog Output frame.

When an Input file is selected, the Output file defaults to the same directory and a name based
upon the Input file name, with an extra letter appended. For example, if the input file was
\masslynx\data\v50.raw the default output file might be \masslynx\data\v50a.raw. When
defaulting the output name, MassLynx attempts to choose a name that doesn’t already exist.

To change the default output file and directory select the Combine Datafile Functions dialog
Output button. The Output File dialog is invoked.

Figure 8.27 Output File dialog

Stopping a Process

To stop a process before it has reached completion, select the Combine Datafile Functions dialog
Stop button. Confirmation of the action will be requested.

The output data file will contain all the information written up to the point at which the process
was stopped.

The Combine All Files Program

General

The Combine All Files program is used to combine a number of files (from the same directory)
that have been acquired using the same acquisition method. Files that have been acquired with the
same method will contain the same number of functions and the same data types for those
functions. There will also be the same number of scans in corresponding functions. The Combine
All Files utility will produce a single output file that will be identical to any one of the input files
in terms of the number of functions, data type, etc.

The combination of the data in this way will result in an increase in the signal to noise ratio shown
by the data.

Page 8-26
 Strip and Combine Functions

Select the MassLynx Tools Shortcut Bar Combine All Files icon to invoke the Combine All Files
dialog. A Menu Bar is provided at the top of the dialog, refer to the “The Combine All Files
Dialog Menu Bar” section, on page 8-29, for details.

Figure 8.28 The Combine All Files dialog

Files to be The controls in this frame display the current MassLynx directory and the
Combined Frame MassLynx data files held within that directory.

Input Files(s) Used to select data files for combination. Files selected for processing
List have a green tick adjacent to them; files not selected have a red cross.
Refer to the “The Combine All Files Dialog Input File(s) List” section, on
page 8-28, for details of how to select and deselect files.

Output Filename Allows the User to enter an output file name. By default, the output
filename is Default.raw. To change the output name enter a new name in
the Output File Name box.

Intensity These options allow the User to specify an intensity threshold for the
Threshold Frame output file. Any peaks in the combined data that fall below the specified
intensity threshold will not be written to the output file. This will help in
controlling the size of the output files (which can approach the size of the
sum of the combined files) by removing peaks that are several orders of
magnitude less intense than the signals of interest.

% full scale Select this option and enter a percentage to set the intensity threshold of
the output file to a percentage of the Base Peak Intensity (BPI).

Intensity Select this option and enter a percentage to set the intensity threshold of
the output file to an absolute intensity.

Page 8-27
Strip and Combine Functions

Peak Intensity
Properties Frame
Mean Peak Sets the intensity of the output file to the mean average of the combined
Intensities peak intensities.

Sum Peak Sets the intensity of the output file to the sum of the combined peak
Intensities intensities.

Peak Separation Data points that fall within the value entered in this text box are combined
Frame together to produce a single peak. For example, if Peak Separation is set
to 2 amu and the mass in question is 200 amu, all peaks between 199 amu
and 201 amu are combined into one peak at 200 amu.

Process button Starts the Combine All Files process.

Note:
This button will change to Cancel when processing has started; this may
be used to stop the process. An output file will almost certainly appear on
the PC’s disk, however it will be incomplete and non-readable by
MassLynx.

Close button Closes the Combine All Files application.

The Combine All Files Dialog Input File(s) List

General

The Combine All Files dialog Input File(s) list (see Figure 8.28) is used to select data files for
combination. Files selected for processing have a green tick adjacent to them; files not selected
have a red cross. Files may be selected/deselected using the mouse and keyboard (see below), or
by using the Menu Bar Operations menu commands, see the “The Combine All Files Dialog Menu
Bar” section, on page 8-29, for details.

Selecting/Deselecting Files using the Mouse and Keyboard

• To select/deselect a single file, double-click on the file name in the Input File(s) list.

• To select/deselect several files, hold down the keyboard Ctrl key while double-clicking on the
required file names.

• To select/deselect a block of files, click on the first file and hold down the Shift key while
clicking on the last file in the block.

Page 8-28
 Strip and Combine Functions

The Combine All Files Dialog Menu Bar

The File Menu

Figure 8.29 The Combine All Files Dialog File menu

Exit Closes the application.

The Operations Menu

Figure 8.30 The Combine All Files Dialog Operations menu

Select Selects the data file currently highlighted in the Combine All Files dialog
Input File(s) list.

Deselect Deselects the data file currently highlighted in the Combine All Files
dialog Input File(s) list.

Select All Selects all the data files in the Combine All Files dialog Input File(s) list.

Deselect All Deselects all the data files in the Combine All Files dialog Input File(s)
list.

Process Starts the Combine All Files process.

Cancel Stops the Combine All Files process.

Note:
If processing is stopped, an output file will almost certainly appear on the
PC’s disk, however it will be incomplete and non-readable by MassLynx.

Processing Files

When all the required files have been selected select the Combine All Files dialog Process push-
button, or select the Menu Bar Operations, Process command. The Process button changes to
Cancel; select this to stop processing.

Note:
If processing is stopped, an output file will almost certainly appear on the PC’s disk, however it
will be incomplete and non-readable by MassLynx.

The field next to the Process/Cancel button displays a progress graphical display, which provides
an indication of the time required to complete the process.

Page 8-29
Strip and Combine Functions

Page 8-30
 Map

Chapter 9 Map

Page 9-1
Map

Contents
Introduction ................................................................................................................................... 9-3
Creating a Data File Map .............................................................................................................. 9-4
To Create a Data File Map ................................................................................................. 9-4
To Stop the Map Process Before it has been Completed ................................................... 9-5
The Map Tool Bar ......................................................................................................................... 9-6
General............................................................................................................................... 9-6
Selecting a Range to Map from the Data File................................................................................ 9-6
Manipulating the Display .............................................................................................................. 9-7
Altering the Display Range with the Mouse ...................................................................... 9-7
Altering the Display Range from the Menu ....................................................................... 9-8
To Change the Map Intensity Scaling ................................................................................ 9-8
Controlling the Appearance of the Display ........................................................................ 9-9
Displaying Diode Array Data........................................................................................... 9-10
Aligning Diode Array Data .............................................................................................. 9-10
Displaying the Chromatogram and Spectrum Windows .................................................. 9-11
The Status Bar.................................................................................................................. 9-11
Selecting the Current Cursor Position .............................................................................. 9-11
User Defined Cursor Positions......................................................................................... 9-13
Editing the Header Information........................................................................................ 9-13
Printing from Map ....................................................................................................................... 9-13
Copying to the Windows Clipboard ............................................................................................ 9-13
General............................................................................................................................. 9-13

Illustrations
Figure 9.1 Typical Map display.................................................................................................... 9-3
Figure 9.2 Map display showing diode array data........................................................................ 9-4
Figure 9.3 The Create Datafile Map dialog .................................................................................. 9-4
Figure 9.4 The Map Data Browser dialog .................................................................................... 9-5
Figure 9.5 The Create Datafile Map dialog .................................................................................. 9-6
Figure 9.6 The Map Display Ranges dialog ................................................................................. 9-8
Figure 9.7 The Map Intensity Scaling Dialog .............................................................................. 9-8
Figure 9.8 The Map View dialog ................................................................................................. 9-9
Figure 9.9 The Diode Array Align dialog .................................................................................. 9-11
Figure 9.10 The Select Mass dialog ........................................................................................... 9-12
Figure 9.11 The Select Wavelength dialog................................................................................. 9-12
Figure 9.12 The Select Time dialog ........................................................................................... 9-12

Page 9-2
 Map

Introduction

Figure 9.1 Typical Map display

The Map program provides a three dimensional representation of an entire data file; this provides
the ability to overview a complete data file very quickly. This is particularly useful for
complicated LCMS data files. The data file can be rapidly searched for particular masses, with the
simultaneous display of mass chromatograms and spectra.

The Map display has three parts. The top trace shows mass chromatogram(s) of the currently
selected mass. The bottom trace shows the spectrum for the currently selected retention time.

Note:
The chromatograms and spectra may be hidden, if required; see the “Controlling the Appearance
of the Display” section, on page 9-9.

Double clicking on the mass chromatogram will invoke the Chromatogram window showing that
mass chromatogram. Double-clicking on the spectrum will invoke the Spectrum window showing
that spectrum.

The middle trace shows the map display of mass against retention time for the data file. The
vertical axis displays mass/charge units (Da/e); the horizontal axis displays retention time in
minutes. The third dimension is the intensity of a particular mass at a particular retention time,
which is represented by a User-selected mapping mode and color scheme.

The currently selected mass and retention time can be changed by moving the cross-hairs cursor
over the display. Moving the cursor in the vertical direction changes the current mass. Moving
the cursor in the horizontal direction changes the current retention time. The current cursor
position is shown on the right hand side of the status bar at the bottom of the display.

For Diode Array data the middle trace shows the map display of wavelength against retention
time. The bottom trace can display either the diode array spectrum at the currently selected
retention time or the mass spectrum at the currently selected retention time.

Page 9-3
Map

Figure 9.2 Map display showing diode array data

In the case of diode array data the vertical axis displays wavelength in nanometers (nm) and the
horizontal axis displays retention time in minutes. The third dimension is the intensity of a
particular wavelength at a particular retention time, which is represented by a User-selected color
scheme.

The current cursor position is given by a pair of cross hairs. At the bottom of the Map window,
the User can display either the diode array spectrum at the currently selected retention time, or the
mass spectrum at the currently selected retention time.

Creating a Data File Map

To Create a Data File Map

1. Select the MassLynx Sample List Menu Bar Map command; the Create Datafile Map dialog
is invoked.

Figure 9.3 The Create Datafile Map dialog

Page 9-4
 Map

2. The data file name displayed in the File: frame is that last used by the program. To change
the data file, select the Browse push-button; the Map Data Browser dialog is invoked.

Figure 9.4 The Map Data Browser dialog

3. Select the required data file.

4. Select the OK push-button; the Map Data Browser dialog is closed.

5. Alter values as required in the Create Datafile Map dialog. If display diode array data is to
be displayed, select the Load Diode Array Data option.

6. Select the OK button. The data file will be read into the Map program and the map display
created. A status bar at the bottom of the map window displays the progress of the Map
process.

To Stop the Map Process Before it has been Completed

In Map, select the Tool Bar button, or select the Menu Bar Process, Stop Process command.

Page 9-5
Map

The Map Tool Bar

General

The Tool Bar can be hidden by selecting the Menu Bar Display, Toolbar command. When the
Tool Bar display is selected, a tick will appear next to it in the Display menu.

Tool Menu equivalent Purpose

Bar
button
File, Open Opens a data file.

File, Print Print the current window in portrait format.

File, Print Print the current window in landscape format.

Edit, Copy Picture Copies the current window to the clipboard.

Process, Stop Process Stops the current map process.

Display, Diode Array Displays a map of the diode array data.

Map
Display, Diode Array Displays the diode array data spectrum.
Spectrum
Display, Scale Edits the intensity scaling for the map display.

Press once to restore the previous display range; press

again to use the default display range.

Selecting a Range to Map from the Data File

By default the Map program will create a map for the whole file, covering the full range of
retention time and mass. To map only the part of the data file, select the Menu Bar Process,
Create Map command; the Create Datafile Map dialog is invoked.

Figure 9.5 The Create Datafile Map dialog

Page 9-6
 Map

Enter values for the Retention Time range and Mass Range that are to be mapped. Reducing the
Retention Time and Mass Range will require less memory and the map process will take less
time. This may be useful for large data files.

It is also possible to reduce the Resolution used for the mass and retention time axes. Reducing
the resolution will reduce memory requirements and may also enhance features in the data.

The Map program will sum all masses in a window equal to the mass resolution to create the map
display. For example, if the mass range is set to 50 amu to 350 amu, and the mass resolution is set
to 1 amu, a point will be plotted at 100 amu, which is a sum of all masses between 99.5 and
100.5 amu.

The Map program will sum all scans in a window equal to the scan resolution to create the map
display. Summing scans in the data file can also improve the signal to noise ratio, this will help to
make peaks more visible and reduce the displayed noise.

Manipulating the Display

Altering the Display Range with the Mouse

General

Mass and retention time axes may both be expanded by clicking on the spectrum. The previous
state of the display can be restored using the Tool Bar button.

Altering the Range of the Retention Time Axis

Press the left mouse button at one end of the region of interest, and without releasing the button,
drag the cursor horizontally to the other end. As the cursor is dragged, a “rubber band” is
stretched out to indicate the range selected; do not go beyond the bounds of the axis. When the
mouse button is released, the selected range will be re-displayed to fill the current window. This
operation can be repeated as often as required.

Altering the Range of the Mass Axis

Press the left mouse button at one end of the region of interest, and without releasing the button,
drag the cursor vertically to the other end. As the cursor is dragged, a “rubber band” is stretched
out to indicate the range selected; do not go beyond the bounds of the axis. When the mouse
button is released, the selected range will be re-displayed to fill the current window.

This operation can be repeated as often as required.

Altering the Range of Both Axes

Press the left mouse button at one end of the region of interest, and without releasing the button,
drag the cursor to the diagonally-opposite corner. As the cursor is dragged, a “rubber box” is
stretched out to indicate the range selected; do not go beyond the bounds of the axis. When the
mouse button is released, the selected region will be re-displayed to fill the current window.

This operation can be repeated as often as required.

Restoring the Display

Pressing the Tool Bar button once restores the display to its previous state. Pressing it a
second time restores the display to the default range.

Page 9-7
Map

Altering the Display Range from the Menu

General

The Map Display menu contains commands for changing the range of the mass axis and restoring
the default display.

To Alter the Range of the Mass axis

1. Select the Menu Bar Display, Range command; the Map Display Ranges dialog is invoked.

Figure 9.6 The Map Display Ranges dialog

2. Enter new Start and End values for the mass and time axes as required.

3. Select the OK button.

Restoring the Display to the Default Range

Select the Menu Bar Display, Default command.

To Change the Map Intensity Scaling

1. Select the Tool Bar button, or select the Menu Bar Display, Scale command; the Map
Intensity Scaling dialog is invoked.

Figure 9.7 The Map Intensity Scaling Dialog

Page 9-8
 Map

2. Set the Intensity Mapping Mode. The options available are Linear, Square Root and Log.
The Log and Square Root intensity modes will give more weighting to lower intensity
masses.

3. Set the Color Palette. The options available are White On Black, Black On White, Gray
Scale, User or one of the Map color schemes. The Map color schemes available are Ocean
Deep, Embers, Emerald Forest, Hot Metal, Cool Metal, Morning Frost, Polar Dawn and
Tropical Lagoon.

The User colors are defined by selecting the MassLynx Tools Shortcut Bar, Colors and
Fonts icon. Select the colors for Data 6 to Data 10 in the Colors and Fonts dialog Type: list
box; refer to the “Changing Colors and Fonts” section, in Chapter 3, “The MassLynx Window
and Related Information” for further details.

4. Set the Map Intensity Range values. Each mass intensity is compared to the most intense
mass in the data file range that is being mapped. Each mass is then mapped according to its
comparative intensity to the corresponding color.

The value of Start (%) corresponds to the percentage intensity at which the color mapping
starts and the value of End (%) corresponds to the percentage intensity at which the color
mapping ends. In the example shown above, all masses with intensities less than 20% on a
logarithmic scale of the most intense mass would be shown in the first User color. All masses
with intensities greater than 80% on a logarithmic scale of the most intense mass would be
shown in the last User color. All masses with intermediate intensities would be mapped to the
other User colors.

5. Select the OK button to exit the dialog and create the map.

Controlling the Appearance of the Display

The appearance of the Map display can be changed using the Map View dialog; this is invoked by
selecting the Menu Bar Display, View command.

Figure 9.8 The Map View dialog

Page 9-9
Map

TIC If this box is checked, the TIC chromatogram of the current data file is
chromatogram displayed at the top of the Map window. Uncheck this control to remove
the TIC chromatogram.

Note:
I f the Menu Bar Display, Diode Array Map option is selected (see the “Displaying Diode Array
Data” section, below), the Map View dialog TIC chromatogram option is replaced by Total
absorbance chromatogram, which operates in the same way.

BPI Displays the BPI chromatogram of the current data file at the top of the
chromatogram Map window. Deselect this option to remove the BPI chromatogram.

Current mass Displays the mass chromatogram of the currently selected mass at the top
chromatogram of the Map window. Deselect this control to remove the mass
chromatogram.

Note:
All chromatograms displayed are overlaid on the same axes.

Current Displays the spectrum at the currently selected retention time at the bottom
spectrum of the Map window. Deselect this control to remove the spectrum.

Cross-hairs Select the color used to display the cross-hairs cursor from the drop down
list box; the settings available are Inverse, Black, White or Axis color.
The cross-hairs cursor can be moved to change the currently selected mass
and retention time.

Map grid, These controls are used to apply a grid to each part of the display. The
Spectrum grid, grid control can be set to Off, Dot, Dash or Solid for each part of the
Chromatogram display.
grid
Automatic link If this option is selected, the Spectrum window will be updated to show
to Spectrum the current spectrum as the cross-hairs cursor is moved across the map
display. Deselect this option to remove the link between Map and
Spectrum.

Automatic link If this option is selected, the Chromatogram window will be updated to
to show the mass chromatogram of the currently selected mass as the cross-
Chromatogram hairs cursor is moved across the map display. Deselect this option to
remove the link between Map and Chromatogram.

Displaying Diode Array Data

The option to display a map of the diode array data is switched on and off by selecting the Tool
Bar button, or by selecting the Menu Bar Display, Diode Array Map option.

The option to display the diode array data spectrum at the bottom of the Map window is switched
on and off by selecting the Tool Bar button, or by selecting the Menu Bar Display, Diode
Array Spectrum option.

Aligning Diode Array Data

Data from the diode array detector may be slightly out of phase with data from the
chromatography system, as there may be a time lag between the sample arriving at the diode array
detector and at the chromatography system.

Page 9-10
 Map

An offset to the time axis of the diode array data can be specified to allow it to manually aligned
with the mass spectral data. Only the display is affected; the data on disk remains unchanged.

To align the diode array data:

1. Select the Menu Bar Display, Diode Array Align command; the Align DAD Time dialog is
invoked.

Figure 9.9 The Diode Array Align dialog

2. Enter the Offset time (mins) that is required to align the data.

3. Select the OK button.

Displaying the Chromatogram and Spectrum Windows

Double clicking on the mass chromatogram will invoke the Chromatogram window for that mass
chromatogram. Double clicking on the spectrum will invoke the Spectrum window for that
spectrum.

The Status Bar

The Status Bar at the Bottom of the Map window displays:

• The current cursor position in terms of mass and retention time.

• The status of an ongoing process such as the Create Map process.

• The function of the currently selected menu item or Tool Bar button.

The Status Bar can be hidden by selecting the Menu Bar Display, Status Bar command; this
toggles the Status Bar off and on. When the Status Bar is selected, a tick will appear next to it in
the Display menu.

Selecting the Current Cursor Position

To Change the Current Cursor Position using the Mouse

Move the mouse cursor to the required position on the Map display and double-click the mouse
button. This position will become the current cursor position. The Spectrum and Chromatogram
displays will be updated accordingly.

If the cross-hairs cursor is displayed, the current cursor position can be changed by clicking
anywhere on the cross-hairs and dragging them to the new position.

To Change the Current Cursor Position from the Menu

1. Select the Menu Bar Display, Select Chromatogram command; the Select Mass dialog (or
Select Wavelength dialog for diode array data) is invoked.

Page 9-11
Map

Figure 9.10 The Select Mass dialog

Figure 9.11 The Select Wavelength dialog

2. Enter the new value in the Current Chromatogram Mass (amu) [or Current Wavelength
(nm) for diode array data] box.

3. Select the OK button.

4. Select the Menu Bar Display, Select Spectrum command; the Select Time dialog is invoked.

Figure 9.12 The Select Time dialog

5. Enter the new value in the Current Spectrum Time (mins) box.

6. Select the OK button.

Page 9-12
 Map

User Defined Cursor Positions

When creating a Map file, MassLynx positions the cross hairs in the center of the display. To
position the cross hairs in a user defined position:

1. Select the Menu Bar Display, Select Chromatogram command; the Select Mass dialog (or
Select Wavelength dialog for diode array data) is invoked, see Figure 9.10.

2. Select the User Defined option; the adjacent text box is enabled, enter a value.

3. Select the OK button.

4. Select the Menu Bar Display, Select Spectrum command; the Select Time dialog is invoked,
see Figure 9.12.

5. Select the User Defined option; the adjacent text box is enabled, enter a value.

6. Select the OK button.

Editing the Header Information

General

The Map Window has a customizable header. Various pieces of information, such as the raw data
file name, can be displayed here, as well as any user text. For more detailed information about the
Header Editor, see the “The Header Editor Dialog” section in Chapter 3, “The MassLynx Window
and Related Information”.

To Change the Displayed Header

1. Select the Menu Bar Display, Header command. The Header Editor (Map Header) dialog
is invoked.

2. Make the required changes.

3. Select the OK button to exit.

Printing from Map

1. Select the Menu Bar File, Print command. The Print dialog is invoked.

2. Make any changes required to the print parameters.

3. Select the OK button to exit and print the Map.

Copying to the Windows Clipboard

General

The Windows Clipboard provides temporary storage for information that is being transferred
between application programs (word processors, spreadsheets, MassLynx etc). A bitmap of the
Map window can be copied to the Clipboard and then, for example, be pasted into a report written
with a Windows compatible word processor.

Page 9-13
Map

To Copy the Map Display to the Clipboard

1. Select the Tool Bar button, or select the Menu Bar Edit, Copy Bitmap command to copy
the contents of the window to the Clipboard.

2. To paste the image into another application, choose the other application’s Edit, Paste
command.

Page 9-14
 Quantify

Chapter 10 Quantify

Page 10-1
Quantify

Contents
Introduction ................................................................................................................................. 10-5
Accessing Quantify ..................................................................................................................... 10-5
MassLynx Automated Quantification - an Overview.................................................................. 10-6
How Does MassLynx Quantify and Report a List of Samples?.................................................. 10-7
Integration of Chromatograms ......................................................................................... 10-7
Generation of Calibration Curves .................................................................................... 10-8
Calculation of Compound Concentrations ....................................................................... 10-9
Displaying Quantify Results .......................................................................................... 10-10
A Step by Step Guide to Quantification .................................................................................... 10-10
1. Create a Sample List ................................................................................................. 10-10
2. Create a Quantify Method......................................................................................... 10-11
3. Start the Analysis ...................................................................................................... 10-20
4. Quantify the Data ...................................................................................................... 10-22
Using the Quantify Window to Examine Results...................................................................... 10-23
General........................................................................................................................... 10-23
The Quantify Window Tool Bar .................................................................................... 10-23
Controlling the Contents of the Quantify Window ........................................................ 10-24
The Quantify Summary Window ................................................................................... 10-25
Selecting the Fields to be Displayed in the Summary Window
and Summary Reports .................................................................................................... 10-26
To Save the Summary Window ..................................................................................... 10-28
The Quantify Graphs Window....................................................................................... 10-28
The Quantify Peak List Window ................................................................................... 10-30
Manually Changing Quantify Results ....................................................................................... 10-32
General........................................................................................................................... 10-32
Manual Peak Integration ................................................................................................ 10-32
To Exclude Erroneous Calibration Points...................................................................... 10-34
To Prevent a Complete Sample from being Used to Create the Calibration Curve ....... 10-35
To Perform Any of the Quantify Processes ................................................................... 10-35
Printing Quantify Reports ......................................................................................................... 10-36
General........................................................................................................................... 10-36
Changing the Printed Quantify Reports Format............................................................. 10-37
Selecting the Chromatogram Display Range for the Quantify Sample Report ......................... 10-39
Copying Quantify Summary Reports to the Clipboard ............................................................. 10-39
Exporting to a LIMS File .......................................................................................................... 10-39
General........................................................................................................................... 10-39
The Header Section........................................................................................................ 10-40
The Samples Section ...................................................................................................... 10-40
The Calibration Section ................................................................................................. 10-41
Files Used During Quantify ...................................................................................................... 10-41
General........................................................................................................................... 10-41
The Sample List (.SPL) File........................................................................................... 10-41
The Quantify Method (.MDB) File................................................................................ 10-42
Peak Lists (.PDB) File ................................................................................................... 10-42
Calibration Curves (.CDB) File ..................................................................................... 10-42

Illustrations
Figure 10.1 The Quantify Manager Bar ..................................................................................... 10-5
Figure 10.2 Chromatogram Integration...................................................................................... 10-7
Figure 10.3 Creating a Compound Calibration Curve................................................................ 10-8
Figure 10.4 Calculating Sample Concentrations ........................................................................ 10-9
Figure 10.5 The Quantify Method Editor dialog...................................................................... 10-12
Figure 10.6 The Secondary dialog ........................................................................................... 10-13

Page 10-2
 Quantify

Figure 10.7 The General Method dialog .................................................................................. 10-15

Figure 10.8 The Integration Window Dialog........................................................................... 10-17
Figure 10.9 The Integrate chromatogram dialog...................................................................... 10-17
Figure 10.10 The Smooth chromatogram dialog ..................................................................... 10-18
Figure 10.11 The Response Threshold dialog.......................................................................... 10-18
Figure 10.12 The Start Sample List Run dialog....................................................................... 10-21
Figure 10.13 The Quantify Samples dialog ............................................................................. 10-22
Figure 10.14 Typical Quantify Window .................................................................................. 10-23
Figure 10.15 The Quantify Display dialog .............................................................................. 10-24
Figure 10.16 Typical Quantify Summary Window.................................................................. 10-25
Figure 10.17 The Report Table Format dialog......................................................................... 10-26
Figure 10.18 Typical Quantify Graphs Window...................................................................... 10-28
Figure 10.19 The Select Entry dialog ...................................................................................... 10-29
Figure 10.20 Typical Quantify Peak List Window .................................................................. 10-30
Figure 10.21 The Format DB List dialog................................................................................. 10-30
Figure 10.22 The List Field Justification dialog ...................................................................... 10-31
Figure 10.23 Typical Chromatogram showing a peak used for calibration point .................... 10-32
Figure 10.24 The Edit Quantify Peak dialog ........................................................................... 10-33
Figure 10.25 The Calibration Curve Edit dialog...................................................................... 10-34
Figure 10.26 The Calibration Curve Edit, Quantify Compounds? dialog................................ 10-34
Figure 10.27 The Quantify Process dialog............................................................................... 10-35
Figure 10.28 The Quantify Reports dialog .............................................................................. 10-36
Figure 10.29 The Print Report dialog ...................................................................................... 10-37
Figure 10.30 The Quantify Report Format dialog.................................................................... 10-38
Figure 10.31 The Quantify Chromatogram Display dialog ..................................................... 10-39

Page 10-3
Quantify

Page 10-4
 Quantify

Introduction
This Chapter is concerned with using MassLynx to perform quantitative assays using the Quantify
application; this is a basic quantitation application, which allows samples to be acquired,
processed and reviewed.

Note:
Quantify is the standard quantitation software supplied with MassLynx. Alternatively, the
QuanLynx Application Manager can be purchased. This provides support for electronic records
and signatures; it is also recommended for some environmental applications. QuanLynx also
includes QuanOptimize, which provides automatic acquisition and quantitation method
development.

The QuanLynx application may be installed as an option when installing MassLynx, see
Chapter 2, “Installing MassLynx”. If QuanLynx is not installed at this stage, Quantify is installed
by default. Refer to the “QuanLynx User’s Guide” for full details of the QuanLynx.

MassLynx enables Quantify calibration curves to be generated, using Standard samples containing
compounds of known concentrations. The calibration curves are then used to calculate the
concentrations of compounds in Analyte samples.

The User provides a list of the samples and a Quantify Method, which describes how to process
each of the compounds within these samples.

The results of Quantify are viewed in the Quantify Summary window. Calibration curves can be
viewed on screen and Quantify Reports can be produced. Quantify information can be copied to
the Windows Clipboard for use by other Windows applications.

The MassLynx automated quantification provides a simple way of quantifying large numbers of
samples within an analysis. Data can be acquired, processed and reports printed without User
intervention. The whole process is controlled from the Sample List Editor.

Accessing Quantify
Quantify is accessed by selecting the MassLynx Bar Quantify tab; this invokes the Quantify
Manager Bar.

Figure 10.1 The Quantify Manager Bar

Page 10-5
Quantify

Available icons are:

Edit Method Invokes the Method Editor dialog, see the “The Quantify Method Editor”
section, on page 10-11.

Process Samples Invokes the Quantify Samples dialog, see the “4. Quantify the Data”
section, on page 10-22.

View Results Invokes the Quantify Window, see the “Using the Quantify Window to
Examine Results” section, on page 10-23 for details.

MassLynx Automated Quantification - an Overview

There are six basic stages involved in automated quantification:

1. Creation of a sample list using the Sample List Editor

2. Acquisition of each sample in the analysis.

3. Integration of data file chromatograms.

4. Generation of Quantify calibration curves.

5. Calculation of compound concentrations.

6. Displaying the results and production of reports.

Page 10-6
 Quantify

How Does MassLynx Quantify and Report a List of

Samples?
After data for all of the samples have been acquired, MassLynx must perform several tasks to
create a printed Report of their concentrations. Whilst the user does have considerable flexibility
in the control of these processes, quantitation is still a straightforward operation, consisting of the
following basic steps:

Integration of Chromatograms

Sample List

Sample 1 Sample 2

Quantification
Method,
All
Compounds

Integrate Integrate Integrate

Chromatograms Chromatograms Chromatograms

Peak List Peak List Peak List

Sample 1
Sample 1 Sample 2 Sample n

Figure 10.2 Chromatogram Integration

The Sample List indicates which sample data files are to be integrated.

Chromatogram integration is made up of two processes: smoothing and peak detection. The
Quantify Method specifies how these are to be applied. Each compound in the Quantify Method
specifies a chromatogram trace that is to be used to Quantify that compound. The chromatogram
for each of the method compounds is integrated and the resulting peaks are saved to a single Peak
List.

Page 10-7
Quantify

Generation of Calibration Curves

Sample List
Calibration Standards

Peak List
Sample 1 Peak List Peak List
Standard 1 Standard 2 Standard n

Quantification
Method,
Compound X

Locate Peak for Locate Peak for Locate Peak for

Compound X Compound X Compound X

Calculate Peak Calculate Peak Calculate Peak

Response Response Response

Fit Curve to
Calibration
Points

Calibration
Curve
Compound X

Figure 10.3 Creating a Compound Calibration Curve

A calibration curve is created for each of the compounds in the Method. Samples, which are to be
used when creating a calibration, are marked as being of type Standard in the Sample List. The
Sample List also specifies the concentration of each of the calibration standards.

The peak, which represents each compound, must be located within a sample’s detected peaks. A
response value for each of the located peaks can then be calculated. For located peaks,
information, such as compound name and peak response, is saved in the Peak List.

Page 10-8
 Quantify

For each compound, one calibration point is obtained from each of the Standard samples.
Calibration points are plotted as response against concentration. A polynomial is fitted to these
points to form the compound’s calibration curve. The calibration curves are saved to a file with
the same name as the Quantify Method.

The Quantify Method specifies how to locate peaks, calculate responses and fit curves.

Calculation of Compound Concentrations

Sample List

Peak List
Sample 1 Peak List Peak List
Sample 1 Sample 2 Sample n

Quantification
Method,
All
Compounds

Locate Peak for Locate Peak for Locate Peak for

All Compounds All Compounds All Compounds

Calculate Peak Calculate Peak Calculate Peak

Responses Responses Responses

Calibration
Curves,
All
Compounds

Apply Apply Apply

Calibration Calibration Calibration
Curves Curves Curves

Compound Compound Compound

Concentrations Concentrations Concentrations
Sample 1 Sample 2 Sample n

Figure 10.4 Calculating Sample Concentrations

Page 10-9
Quantify

MassLynx calculates the concentration of each of the Method compounds for the samples in the
Sample List.

The peak, which represents each compound, must be located within a samples Peak List. A
response value for each of the located peaks can then be calculated. For located peaks,
information, such as compound name and peak response, is saved in the Peak List.

A concentration is calculated for each of a compound’s located peaks by applying the compound’s
calibration curve. Concentration information is saved in the Peak List.

Displaying Quantify Results

The Quantify Window is used to display and report the results of quantification; see the “Using
the Quantify Window to Examine Results” section, on page 10-23 for details.

A Step by Step Guide to Quantification

1. Create a Sample List

General

A list of samples to be used to perform the analysis must be first created using MassLynx, see
Chapter 4, “Sample Lists”. These samples can be acquired manually, but more often they are
acquired automatically using an autosampler. The Sample List Editor is part of the MassLynx
top-level screen; it has user-selectable columns, e.g. File Name, Bottle Number and Sample
Type, into which the appropriate information for each sample is entered. Each sample is
displayed as a single row in the Sample List.

To enable MassLynx to perform a complete analysis, the following must be described:

• The Sample Type for each bottle in the autosampler, i.e. whether it is a standard, an analyte, a
blank, or a Quality Control (QC) sample.

• How the sample is to be acquired.

• If the sample is a standard or QC, its concentration(s).

In addition, MassLynx must be given a file name in which to store the data. Management
information, such as Sample ID, the submitter’s name, or a sample description may also be
specified.

Projects

MassLynx allows the work to be organized in a Project, which is a simple way of organizing all
the data files, methods and results for a particular assay into one directory structure. When a
MassLynx Project is opened, a new directory is created to hold all the files associated with the
Project.

The types of file that can be saved in a MassLynx Project are:

• Raw data files.

• QuanLynx Datasets.

• Sample Lists.

Page 10-10
 Quantify

• Quantify Methods.

• Quantify calibration curves.

• Tuning files.

• Scan methods.

• Instrument calibration files.

• Inlet methods.

Projects are created and selected from the MassLynx menu bar File command. See Chapter 3,
“The MassLynx Window and Related Information” for details of how to create, or open, a Project.

2. Create a Quantify Method

General

A Quantify Method must be created before Integration or Quantification can be performed.

The Quantify method describes how a data file is processed to produce calibration curves and
quantitative information. Details must be entered into the method for each of the compounds
being used in the analysis.

The Quantify Method specifies information for performing the following tasks:

• Integration of a chromatogram trace to obtain peak information.

• Location of the chromatogram peak relating to a specific compound from the list of detected
peaks.

• Calculation of a response factor for the located peak.

• Formation of a Quantify calibration curve.

The Quantify Method Editor

The Quantify Method Editor dialog creates new methods and modifies existing ones; it is
invoked by selecting the Quantify Manager Bar Edit Method icon.

When invoked, the Quantify Method Editor contains the current MassLynx Method; if this is not
available, the Editor will contain default values and the name of the current Method in the
Method Editor title bar is set to [Untitled].

The current Quantify Method Editor Method is the current system Method file; it is used when
performing quantitation.

Note:
Changes made to the Method are not made permanent until they have been saved to disk.
Consequently, the Method must be saved before it can be used to perform quantitation; select the
Method Editor Menu Bar File, Save command to update the current Method file, or File, Save
As to save to a new Method file.

Page 10-11
Quantify

Figure 10.5 The Quantify Method Editor dialog

Setting the Quantify Method Parameters

1. Enter the name of the compound in the Quantify Method Editor, Name box. (This can be up
to 40 characters in length.)

2. Select the appropriate button to add the compound to the list in the Compound: box:

Append Adds the compound to the end of the list.

Insert Inserts the compound before the currently selected compound in the list.

Modify Changes the currently selected compound to that entered in the Name box.

Delete Deletes the currently selected compound from the list.

3. Select the internal reference compound in the Internal Ref box. (Set this to [None] if the
compound is not using an internal reference. Only compounds that appear in the compound
list can be selected.)

Note:
The Multi button invokes the Multiple Internal Standards dialog; this dialog is valid only
in QuanLynx, it is not relevant to Quantify.

4. Set the Quantify Trace edit control to the trace descriptor of the chromatogram being used to
quantify the compound. (Quantify Trace specifies a chromatogram to be integrated when
performing automatic peak detection and is used during the locate phase when matching Peak
List entries against Method compounds). The trace descriptor should be:

• A single decimal number for mass chromatograms.

• Two decimal numbers separated by a “>” for a Multi Reaction Monitoring (MRM)
function, e.g. 274.10 > 182.10. The first number represents the parent mass; the second
number represents the daughter mass. A reaction can be specified for MRM data. The
chromatogram used can be constructed from multiple chromatograms using the Add (+),
Subtract (-) and Range (:) operators.

Page 10-12
 Quantify

• TIC for Total Ion Current chromatograms.

• BPI for Base Peak Intensity chromatograms.

• An1, An2, An3 or An4 for analog data, depending on the channel required.

• The wavelength for Diode Array Detector (DAD) data.

• Ch1, Ch2, etc. for Selective Ion Recording (SIR) data to use one Quantify Method with
multiple SIR functions, where Ch1 is the first mass in the list, Ch2 is the second, etc.

Note:
The trace descriptor value will be entered automatically in Quantify Trace edit control if the
Peak Location parameters are entered using the mouse, see Step 8.

5. In specific cases, it may be necessary to specify a secondary ion; if so, select the Sec>>
button; the Secondary dialog is invoked.

Figure 10.6 The Secondary dialog

• Enter the mass of the secondary ion in the Trace field. (If this field is left blank, the
secondary ion will not be used during peak location.)

• Enter the expected ratio between the size of the Primary and Secondary peaks in the
Expected Primary/Secondary ratio field. (If this field is set to zero, the peak ratio will
not be used for compound location.)

• Select the OK button to accept the new settings; the display returns to the Quantify
Method Editor.

6. For multifunction data, select which function number is to be used to quantify the current
compound in the Acquisition Function Number list box. (Any number between One and
Thirty-two, or Any, may be selected.)

7. Set the Concentration of Standards box to the Sample List column that contains the
compound’s concentration level within each Standard or QC sample; e.g. Conc A if the
concentration is defined in the CONC_A column in the Sample List. (The software allows up
to twenty concentration levels within a single sample. If the compound is an Internal
Standard and is at the same concentration in all samples, the Fixed option can be selected.
The parameter box adjacent to the Concentration of Standards box becomes active when
the Fixed option is selected; enter the concentration level of the International Standard in the
box.)

8. Select the Peak Location Method by selecting either the Retention Time (mins) or Relative
Retention Time option in the Peak Location frame. Alternatively, a method compound to
use as the retention time reference can be selected from the RT Ref drop-down list. (The
Peak Location Method determines how a peak within a Peak List is identified as matching a
method compound. If a reference is entered, the expected retention time of the compound

Page 10-13
Quantify

will be shifted by the same amount as the found reference peak from its predicted time.)
Selecting the Zero button will zero all the retention times.

Note:
The Peak Location Time Window (mins) ± (see below,) and Retention Time (mins) or
Relative Retention Time parameter values can be entered by using the mouse, or with the
keyboard.

To use the mouse, proceed as follows:

• Arrange the MassLynx display so that the Quantify Method Editor and the
Chromatogram window, showing the chromatogram to be used, are both visible. Select
the Compound for which parameters are to be set in the Method Editor.

• On the Chromatogram window, press the right-hand mouse button at one end of the
chromatogram region of interest and, without releasing the button, drag horizontally to
the other end. As the mouse is dragged, a “rubber band” will be displayed to indicate the
range selected. The Quantify Method Editor window will be updated to show the new
Peak Location Time Window (mins) ±, and the Retention Time (mins) or Relative
Retention Time will be set to the middle point of the Time Window (mins) ±.

The Quantify Trace parameter will be set to the same type as the selected chromatogram [i.e.
Total Ion Count (TIC), Base Peak Intensity (BPI), mass chromatogram or Multiple reaction
Monitoring (MRM)].

• The Retention Time (mins) or Relative Retention Time parameters can also be set with
a single click of the right-hand mouse button on the chromatogram trace.

To use the keyboard, proceed as follows:

• If Retention Time (mins) has been selected, set it to the time, in decimal minutes, at
which the compound is expected to elute. Set the Peak Location Time Window (mins) ±
parameter to specify by how much the compound elution time may vary. [The Time
Window (mins) ± is applied either side of the predicted retention time to give a valid
window. The Time Window (mins) ± parameter, multiplied by the factor entered in the
Integrate chromatogram dialog (see the “Setting Quantify Method Peak Integration
Parameters” section, on page 10-16), defines the chromatogram range that will be
integrated.]

• If Relative Retention Time has been selected, set it to the time at which the compound is
expected to elute relative to the compound specified in the Internal Ref control. (The
value specified here is a multiplication factor that is applied to the time at which the
internal reference compound elutes. This can be used to deal with situations where some
drift may occur in the time at which compounds elute, but their relative retention times
remain constant.)

9. Set the Peak Selection parameter to specify which peak should be located when more than
one peak is detected within the Peak Location Time Window. (By default, the peak Nearest
to the specified retention time will be selected. Other options that can be selected are Largest
peak, First peak or Last peak in the specified time window, and Totals. Totals allows the
sum of the valid peaks within the window to be calculated.)

10. If required, set the User Peak Factor. (This value is a multiplication factor that will be
applied to all calculated concentrations for the current compound. If the User Peak Factor is
left at 0, or set to 1, the concentration values will not be changed.)

Page 10-14
 Quantify

11. If required, select the User RF Value option and enter a Response Factor (RF) value in the
control. (The User RF Value is used in cases where there are no calibration standards to plot
a calibration curve. It represents the gradient of a curve and is used as a multiplication factor,
which will be applied to peak responses for the current compound to determine
concentrations.)

12. Select the General Parameters button; the General Method dialog is invoked. (To use
these General Parameters for all compounds in the Method, choose the Quantify Method
Editor Menu Bar Edit, Propagate, General Parameters command. A tick mark will appear
next to this option and the General Parameters will be copied to all compounds in the
Method.)

Figure 10.7 The General Method dialog

The Response frame parameters determine how the response value of a located peak is to be
calculated. The response values are used to form calibration curves for compounds from
standard samples, and to calculate the concentration of compounds within analyte samples.

13. Select the Response frame, Type option:

Internal Should be selected if a compound’s response is to be calculated using an

(relative) Internal Standard, in which case the Quantify Method Editor, Internal
Ref control must have the Internal Standard compound selected.

External Should be selected if the compound does not have an Internal Standard;
(absolute) the response is then taken as the absolute peak height/area.

14. Select the Response frame, Areas or Heights option to specify whether the compound
responses will be based on peak areas, or heights, respectively.

15. Set the Calibration Curves frame parameters, as shown in Steps 16 to 19. (The Calibration
Curve parameters determine how a compound’s calibration curve is to be created.)

16. Select the type of calibration curve in the Polynomial Type list box; the following options are
available:

Average RF Produces a calibration that is a straight line through the origin and through
the mean response factor of the calibration points. A response factor is the
response of a calibration point divided by its concentration. This option
should be selected for compounds where the Method Editor,
Concentration of Standards list box is set to the Fixed option.

Page 10-15
Quantify

Linear Performs a linear regression on the compound’s calibration points.

Quadratic Performs a second order regression on the compound’s calibration points.

Cubic Performs a third order regression on the compound’s calibration points.

Quartic Performs a fourth order regression on the compound’s calibration points.

17. Set the Point of Origin option to Exclude, Include or Force. (At the point of origin, it is
assumed that zero concentration has a response of zero. If Polynomial Type is set to
Average RF this parameter is not used and the option is grayed out.)

Force The calibration curve will always pass through the origin.

Include The point of origin will be included in the calibration curve regression; the
curve will not usually pass through the origin.

Exclude The origin will be ignored when forming the calibration curve.

18. Set the Fit Weighting option to None, 1/X, 1/X^2, 1/Y, or 1/Y^2. (This parameter is used to
give higher priority to calibration points with a low concentration or response when using
regression to fit a calibration curve. This generally results in the calibration curve being fitted
closer to points at low concentrations, hence reducing the relative error at these points. If
Polynomial Type is set to Average RF this parameter is not used and the option is grayed
out.)

19. Set the Axis Transformation parameter to the required option. (The available options are
None, Ln (Natural Log), Log (Base 10 Log) and Square Root. The transformation is applied
to the concentration and response values before the calibration curve is fitted. This option is
only available when Point of Origin is set to Exclude, and Fit Weighting is set to None.)

Note:
Axis transformations cannot be used with RF-type curves, curves that use point weighting, or
curves that include, or force, the origin.

20. If required, set the Concentration frame Units parameter. (The value set here will be used
on the concentration axis of calibration curves, and in the concentration column header in the
Summary Report.)

21. Select the OK button to accept the new settings; the display returns to the Quantify Method
Editor.

Setting Quantify Method Peak Integration Parameters

The Peak Integration parameters are used when automated chromatogram peak detection is being
performed. The integration parameters can either be set on a per compound basis or for all
compounds within the method.

The facility to set different integration parameters for different compounds can be useful where
peak characteristics, such as peak width or shape, vary between different compounds.

To use the same integration parameters for all compounds in the Method, select the Quantify
Method Editor Edit, Propagate, Integrate Parameters command. A tick mark will appear next
to this command and the integration parameters will be copied to all compounds in the Method.

By default, integration will take place over the chromatogram range defined by the Quantify
Method Editor, Time Window (mins) ± parameter. To integrate over a larger window, select
the Quantify Method Editor Menu Bar Edit, Integrate Window command; the Integration

Page 10-16
 Quantify

Window dialog is invoked. Enter a multiplication factor in the Factor applied to location
window to calculate integration window text box. This factor will be applied to the Peak
Location Time Window to calculate the integration window; it is the same for all compounds in
the Method.

Figure 10.8 The Integration Window Dialog

To define the integration parameters, select the Quantify Method Editor, Integrate Parameters
button; the Integrate chromatogram dialog is invoked.

Figure 10.9 The Integrate chromatogram dialog

The peak-to-peak noise amplitude value is entered in the Peak-to-peak amplitude text box; the
integration software uses this to pre-filter the chromatogram. A suitable value can be measured
directly from a chromatogram by pressing the right-hand mouse button, and dragging the mouse
across a section of noise in the chromatogram. The sensitivity of the integration algorithm can be
fine-tuned by manually adjusting this value.

The Copy and Paste buttons allow integration parameters to be written to, and read from, the
Windows Clipboard. This enables integration parameters to be transferred easily between the
chromatogram and the Quantify Method. This can be useful when experimenting to find the
correct integration parameters using a chromatogram.

Select the ApexTrack Peak Integration option to use ApexTrack peak detection algorithm, see
Chapter 6, “Chromatogram” for details.

Smoothing

The chromatogram may be smoothed, before integrating, by selecting the Integrate

chromatogram dialog Enable smoothing option. The parameters for the smooth may be
examined and altered by selecting the Smooth button; this invokes the Smooth chromatogram
dialog.

Page 10-17
Quantify

Figure 10.10 The Smooth chromatogram dialog

The Window size (scans) ± parameter should be set to the half-width of the smoothing window in
scans. This parameter can be set automatically by clicking the right-hand mouse button, and
dragging across a chromatogram peak.

Set the number of times the smooth is repeated, by changing the Number of smooths parameter
from its default value of two. Increasing this parameter gives a heavier smooth.

Two types of smoothing are available for chromatograms; Moving Mean and Savitzky Golay.
Both methods slide a window along the chromatogram, averaging the data points in the window to
produce a point in the smoothed spectrum. Moving Mean takes the arithmetical mean of the
intensities of the data points in the window. Savitzky Golay takes an average of the intensities
weighted by a quadratic curve. This tends to enhance peak and valley shapes, as well as
preserving the height of the peaks better than the Moving Mean. However, Savitzky Golay does
tend to produce small artifacts on either side of the real peaks.

Once the parameters have been selected, select the OK button to return to the Integrate
chromatogram dialog.

Peak Thresholding

Small peaks may be optionally removed by setting one of the four available threshold parameters.
To examine or modify these parameters, select the Integrate chromatogram dialog Threshold
push-button; the Response Threshold dialog is invoked.

Figure 10.11 The Response Threshold dialog

Relative height Removes the peaks whose height is less than the specified percentage of
the highest peak.

Absolute height Removes the peaks whose height is less than the specified value.

Relative area Removes the peaks whose area is less than the specified percentage of the
largest peak area.

Absolute area Removes the peaks whose area is less than the specified value.

Once the parameters have been selected, select the OK button to return to the Integrate
chromatogram dialog.

Page 10-18
 Quantify

Peak Detection

The parameters controlling the positioning of peak baselines may be examined and modified by
selecting the Integrate chromatogram dialog Peak detect button. The dialog invoked, and
corresponding parameters, will depend on the Peak Detection method previously selected by the
Integrate chromatogram dialog ApexTrack Peak Integration option.

Refer to Chapter 6, “Chromatogram” for full details of Peak Detection.

Creating a New Quantify Method

1. Select the Quantify Method Editor dialog File, New command. (The Quantify Method
Editor controls are set to default values and the Compound: list box is emptied. The name
of the current Method in the Quantify Method Editor title bar is set to [Untitled].)

2. Add the desired compounds as described below.

3. Select the Menu Bar File, Save As command; the Save As dialog is invoked.

4. Enter the name of the new Method into the Save As dialog.

5. Select the OK button.

Selecting an Existing Quantify Method

1. Select the Quantify Method Editor Menu Bar File, Open command; the Open dialog is
invoked.

2. Choose the required Method file from the Open dialog.

3. Select the Open button. (The compounds held within the Method are loaded into the
Quantify Method Editor Compound: list box. The first compound within the Method is
selected.)

To Propagate General Parameters to All Compounds in the Quantify Method

To use the same general parameters for all compounds in the method, select the Quantify Method
Editor Edit, Propagate, General Parameters command. (A tick mark will appear next to this
command and the general parameters will be copied to all compounds in the method.)

To Propagate Integration Parameters to All Compounds in the Quantify Method

To use the same integration parameters for all compounds in the method select the Quantify
Method Editor Menu Bar Edit, Propagate, Integrate Parameters command. (A tick mark will
appear next to this command and the integration parameters will be copied to all compounds in the
method.)

To Add a New Compound to the Quantify Method

1. Enter the required information for the new compound in the Quantify Method Editor.

2. Select the Append button; the new compound will be added to the end of the Compound:
list.

To Insert a New Compound in the Quantify Method

1. Select the entry in the Quantify Method Editor, Compound: list before which the new
compound is to be inserted.

Page 10-19
Quantify

2. Enter the required information for the new compound.

3. Select the Insert button, the new compound will be inserted in the Compound: list.

To Modify Information for an Existing Compound in the Quantify Method

1. Select the entry in the Quantify Method Editor Compound: list that is to be modified.

2. Enter the updated information.

3. Select the Modify button.

To Delete a Compound from the Quantify Method

1. Select the entry in the Quantify Method Editor Compound: list that is to be deleted.

2. Select the Delete button, or the Delete key.

To Delete all Compounds in the Method

1. Choose the Quantify Method Editor Edit, Delete All Compounds command, the Method
Editor, Delete all entries dialog is invoked.

2. Select the OK button to delete all compounds in the method.

3. Start the Analysis

Before starting an analysis, save any changes made to the MassLynx Sample List by selecting the
MassLynx File, Save command.

To begin acquiring data select the MassLynx Menu Bar Run, Start command, or select the Tool

Bar button; this invokes the Start Sample List Run dialog.

Project The name of the current project. To acquire to a different project, exit this
dialog, open another project, and start acquisition again.

Acquire Sample Acquires data for all the samples in the Sample List. See the appropriate
Data Instrument User's Guide for further details.

Auto Process Processes the acquired data as specified in the Process column of the
Samples Sample List.

Auto Quantify Quantifies the acquired data using the method specified in the Quantify
Samples Samples dialog (see below). If a method is not defined in the Quantify
Samples dialog, the current method will be used. If selected, this option
will generate a QuanLynx Dataset with the same name as the Sample List;
if a Dataset of this name already exists, a numeric postfix will be
appended to the name. The Dataset can be viewed in the QuanLynx
Browser, which is invoked by the Quantify Application Bar, View
Results icon.

Note:
The above three actions can be run together or independently, i.e. data can be acquired,
processed and quantified in one go, or acquired in one run and processed or quantified it at a
later date.

Page 10-20
 Quantify

Figure 10.12 The Start Sample List Run dialog

Run Frame

From Sets the range of samples in the Sample List that will be acquired/and or
Sample… analyzed.
To Sample
Priority Marks this entry as a Priority process.

Note:
The MassLynx Queue Properties dialog Pre-emptive Scheduling option must be selected.

Night Time Select this option to mark this entry as a night time process.
Process
Note:
The MassLynx Queue Properties dialog Night Time Scheduling option must be selected.

Process

Pre-Run, When submitting a batch to the MassLynx Queue to be acquired, processed,

Post-Run etc., these fields allow the User to specify an executable to be run before the
batch starts and when the batch has finished. Any .EXE file can be run,
hence this allows Users to write their own applications to perform some
task before, or after, a batch is executed

Page 10-21
Quantify

4. Quantify the Data

Once data has been acquired, it can be quantified by creating a Dataset containing the samples
specified in the Sample List. Select the Quantify Application Bar, Process Samples icon to
invoke the Quantify Samples dialog. Select the options as required and select the OK button.

Figure 10.13 The Quantify Samples dialog

Integrate Integrates all the sample data files named in the Sample List.
Samples
Calibrate Uses Integration results to create Quantify calibration curves. Do not
Standards select this option if an existing calibration is to be used; in this case use the
Curve:, Browse button to select the desired calibration file

Quantify Uses Integration results and Quantify calibration curves to calculate

Samples compound concentrations.

Print Quantify Produces hard copies of the results of integration and quantitation.
Reports
Export Results Produces a text file containing the quantitation results details for use with
to LIMS LIMS systems. If this option is selected, the LIMS Export Frame File:
Browse button is enabled; select the Browse button and select a file, or
enter the name of a new one, and select the Save button.

Project The name of the current project. To quantify using a different project, exit
this dialog, change the current project and select the MassLynx Quantify
Application Bar, Process Samples icon again.

Quantify Frame

From Sets the range of samples in the Sample List that will be quantified.
Sample…
To Sample
Method: or To change the files select the appropriate Browse button and select a new
Curve: file.

Page 10-22
 Quantify

Using the Quantify Window to Examine Results

General

The Quantify Window is invoked by selecting the MassLynx Quantify Application Bar, View
Results icon.

Figure 10.14 Typical Quantify Window

The Quantify Window has a Menu Bar and Tool Bar; it uses a Multiple Document Interface
(MDI) display which allows multiple windows to be displayed simultaneously.

There are three windows; the Summary Window, the Graphs Window and the Peak List
Window.

The Quantify Window Tool Bar

The Tool Bar is displayed at the top of the Quantify Window.

Tool Menu equivalent Purpose

Bar
button
File, Print Prints the current Window in portrait format.

File, Print Prints the current Window in landscape format.

Shows the previous peak in the Summary Window.

Page 10-23
Quantify

Tool Menu equivalent Purpose

Bar
button
Shows the current peak in the Summary Window.

Shows the next peak in the Summary Window.

Window, Tile Displays the current windows in a tiled view.

Window, Cascade Displays the current windows in a cascaded view.

Window, Stack Displays the current windows in a stack view.

Selects the current entry.

Decrements the current entry in the Summary Window.

Increments the current entry in the Summary Window.

Views the previous Sample Group. This applies to data

acquired by QuanLynx.

Views the next Sample Group. This applies to data

acquired by QuanLynx.

Press once to restore the default display range.

Controlling the Contents of the Quantify Window

Select the Menu Bar Display, View command; the Quantify Display dialog is invoked.

Figure 10.15 The Quantify Display dialog

Graphs Frame

Curve Displays the calibration curves.

Page 10-24
 Quantify

Residuals Displays the residuals.

Edit Header Invokes the Header Editor dialog; this edits the document header
displayed at the top of the Graphs window. See Chapter 3, “The
MassLynx Window and Related Information” for further information.

Summary Frame

Show Displays the Summary Window.

Summary
Window
Format Select the required Summary Window Format from the list box. The
options are List By Compound and List By Sample.

Peak List Frame

Show Window Displays the Peak List Window.

Edit Header Invokes the Header Editor dialog; this edits the document header
displayed at the top of the Peak List window. See Chapter 3, “The
MassLynx Window and Related Information” for further information.

The Quantify Summary Window

Figure 10.16 Typical Quantify Summary Window

Page 10-25
Quantify

The Quantify Summary Window displays a summary of the results of quantification. The results
can be listed by compound or by sample. If no peak has been located for a compound entry, the
peak information fields will be left blank.

The Quantify Window Tool Bar buttons can be used to display information about a new
compound/sample.

The format of the Quantify Summary Window also determines the format of the printable
Summary Reports. Two types of Summary Reports can be printed:

• Listed by sample.

• Listed by compound.

Many different columns of quantification information can be displayed in the Summary Window;
the User can select the columns to display. Use the horizontal and vertical scroll bars, if available,
to move around the Summary Window display.

Selecting the Fields to be Displayed in the Summary Window and

Summary Reports

General

The formats of the Summary Window, when listed by sample and when listed by compound, are
changed independently. Double-click on one of the Summary Window column headers, or select
the Quantify Summary Window Menu Bar Edit, Output Compound Format command, or
Output Sample Format command, as appropriate; this invokes the Compound Report Table
Format dialog, or Sample Report Table Format dialog. These dialogs are identical in function.

Figure 10.17 The Report Table Format dialog

The fields currently present in the Summary document are shown in the Displayed Fields: list.
Fields that can be added to the Summary document are shown in the Available Fields: list.

Page 10-26
 Quantify

Any changes made here will be reflected in the Summary Window and Summary Reports. The
Summary Reports will show the maximum number of columns that will fit on one page. To
include more columns print in landscape mode instead of portrait mode.

To Append New Fields in the Summary Window

1. Highlight the required field in the Available Fields: list box.

2. Select the Append button. The field is appended to the list in the Displayed Fields: box.

3. Repeat steps 1 and 2, as required.

4. Select the OK push-button to save the changes and exit.

To Insert New Fields in the Summary Window

1. Highlight the required field in the Available Fields: list box.

2. Highlight the field before which the new field is to be inserted in the Displayed Fields: list
box.

3. Select the Insert button. The field is inserted in the list in the Displayed Fields: box.

4. Repeat steps 1 to 3, as required.

5. Select the OK button to save the changes and exit.

To Remove a Field From the Summary Window

1. Highlight the field to be removed in the Displayed Fields: list box.

2. Select the Remove push-button. The field is removed from the Displayed Fields: list box.
To remove all the fields in the Summary Window select the Remove All push-button.

3. Repeat steps 1 to 2, as required.

4. Select the OK push-button to save the changes and exit.

To Format the Field Display in the Summary Window

1. Highlight the field whose display settings are to be altered in either the Available Fields: or
Displayed Fields: list boxes.

2. Enter the required header text for this field in the Header text box.

3. Change the Justification setting to LEFT, RIGHT, or CENTRE, as required.

4. Change the Field Width and Decimal Places, as required.

5. Change the setting of the Not Found control as required. The Not Found control determines
what will be printed in the Summary Report for this field, if the peak is not found. The
options are Blank, Zero, Dash, Not found or n/a.

6. Repeat steps 1 to 5, as required.

7. To change the settings for all fields back to the default values, select the Default push-button.

8. Select the OK push-button to save the changes and exit.

Page 10-27
Quantify

To Save the Summary Window

Select the Summary Window Menu Bar File, Save Summary by Compound or Save Summary
by Sample command, as appropriate.

The Quantify Graphs Window

General

Figure 10.18 Typical Quantify Graphs Window

The Quantify Graphs Window contains a graphical display of the current calibration curve and/or
its residuals plot. Statistical information on the calibration curve is displayed above the graphs. A
User-configurable document header may be displayed at the top of the Window.

The current calibration curve file holds a calibration curve for each of the compounds being
analyzed. Other calibration curves to be easily selected by selecting the Tool Bar and
buttons.

The calibration curve graph displays concentration against response value. The vertical axis is
labeled as a percentage of the maximum response. The horizontal axis is labeled with the
concentration units specified in the method. The displayed calibration curve shows the response
value expected for particular concentrations. Crosses mark the calibration points used to create
the curve.

The residual plot displays concentration against delta concentration at the calibration points. This
shows the difference between the concentration predicted by the calibration curve and the actual
concentration at the calibration points.

Page 10-28
 Quantify

Selecting a Different Calibration Curve

To select another calibration curve, from within the current file, using the Tool Bar:

Select the button to show the previous calibration curve.

Select the button to show the next calibration curve.

Select the button to invoke the Select Entry dialog; this allows the number of the required
calibration curve to be entered. Curve number 1 is for the first compound, curve number 2 the
second, etc.

Figure 10.19 The Select Entry dialog

Changing the Calibration Curve Display Range

Both the horizontal and vertical display ranges of the Graphs Window can be expanded. Press the
mouse button at one end of the region of interest, and without releasing the button, drag the mouse
horizontally or vertically or in both directions to the other end. As the mouse is dragged, a
“rubber band” is stretched out to indicate the selected range; don’t go beyond the bounds of the
axis. When the mouse button is released the selected range will be re displayed to fill the current
window.

This operation can be repeated as often as required.

Selecting the Tool Bar button restores the display to the default range.

Changing the Current Calibration Curve File

To view another calibration curve file, select the Menu Bar File, Calibration command; the File
Open dialog is invoked. Select a file from the list box and select the Open push-button.

Displaying More Information about a Particular Calibration Point

Click on a calibration point to update the Summary and Peak List Windows to display that
calibration point as the current entry.

Double-clicking on a calibration point displays the Peak List entry and invokes the corresponding
chromatogram. The Edit Quantify Peak dialog is also automatically invoked allowing the User
to make manual adjustments to the baseline assignment. A comment can also be stored in the
Peak List for this particular peak. For more information see “Manual Peak Integration” section on
page 10-32.

Page 10-29
Quantify

The Quantify Peak List Window

General

Figure 10.20 Typical Quantify Peak List Window

The Quantify Peak List Window contains a text listing of all the peaks within the current Peak
List; the current peak is highlighted. Displayed Peak List columns are User-configurable. Use the
horizontal and vertical scroll bars, if available, to move around the Peak List display.

A User-configurable document header can be displayed at the top of the Peak List window.

Configuring Displayed Peak List Columns

The Peak List Window allows all the information from a Peak List entry to be displayed. It is
possible to select which columns are to be displayed and in which order they are to appear.

Double-click on one of the Peak List Window column headers, or select the Peak List Window
Menu Bar Display, PeakList display format command; this invokes the Format DB List dialog.

Figure 10.21 The Format DB List dialog

To Append New Fields in the Peak List Window

1. Highlight the required field in the Fields list box.

2. Select the Append button. The field is appended to the list in the Format box.

3. Repeat steps 1 and 2 as required.

4. Select the OK push-button to save the changes and exit.

Page 10-30
 Quantify

To Insert New Fields in the Peak List Window

1. Highlight the required field in the Fields list box.

2. Highlight the field before which the new field is to be inserted in the Format list box.

3. Select the Insert button. The field is inserted in the list in the Format box.

4. Repeat steps 1 to 3 as required.

5. Select the OK button to save the changes and exit.

To Remove a Field From the Peak List Window

1. Highlight the field to be removed in the Format list box.

2. Select the Remove push-button. The field is removed from the Format list box. To remove
all the fields in the Peak List Window select the Remove All push-button.

3. Repeat steps 1 to 2, as required.

4. Select the OK push-button to save the changes and exit.

To Format the Field Display in the Peak List Window

1. Highlight the field whose display settings are to be altered in either the Fields or Format list
box.

2. Select the Justification push-button. The List Field Justification dialog is invoked.

Figure 10.22 The List Field Justification dialog

3. Select the required heading for the field in the Field Name list box.

4. Change the Justification setting to Left, Centre, or Right, as required.

5. Enter the required values in the Width:, SF: (Significant Figures) and DP: (Decimal Places)
text boxes.

6. Select the OK push-button to save the changes and exit.

Changing the Current Peak List File

To view another Peak List file, select the Menu Bar File, Peak List command; the File Open
dialog is invoked. Select a file from the list box and select the Open push-button.

Page 10-31
Quantify

Displaying Peak List Chromatograms

To display the chromatogram and peak associated with a Peak List entry, double-click on the
required entry.

Manually Changing Quantify Results

General

Although MassLynx can perform a complete automated quantification analysis from setting up a
Sample List and acquiring data to printing Quantify Reports, it is also possible to repeat individual
Quantify processes and to manually edit results including:

• Manual editing of peak baselines.

• Editing calibration curves to exclude erroneous calibration points.

• Performing Quantify Locate compounds, Calculate calibration curves or Quantify

compounds processes.

Manual Peak Integration

If the automated peak detection is not determining peak baselines satisfactorily it is possible to
define the baselines manually. This can be achieved by modifying the peak information held in
the Peak Lists or by creating them from scratch.

To display an integrated peak in Chromatogram, double-click on the required entry in the

Summary Window or the Peak List Window. Calibration standard peaks can be selected by
double-clicking on the desired calibration point in the Calibration Curve Window.

The Chromatogram Window will be invoked showing the relevant peak. Also, the Edit Quantify
Peak dialog is automatically invoked, allowing the User to make manual adjustments to the
baseline assignment.

Figure 10.23 Typical Chromatogram showing a peak used for calibration point

Page 10-32
 Quantify

Figure 10.24 The Edit Quantify Peak dialog

The peak baseline can be modified by clicking on, and dragging, the handles that appear at either
end of the baseline. The information in the Edit Quantify Peak dialog Peak Information Frame
is updated. When satisfied with the manual integration, select the OK push-button to save the
new peak integration information. A Comment can also be stored in the Peak List for this
particular peak; this comment can be included in the printed report.

If no peak was detected, the chromatogram trace, which should have contained the peak, can be
displayed by double-clicking on the appropriate Summary Window entry. A baseline can be
added by pressing the right mouse button at one end of the chromatogram region of interest, and
dragging the mouse horizontally to the other end. A “rubber band” is stretched out to indicate the
range selected, and a baseline will be drawn.

To delete the current peak, select the Delete button, followed by the OK button.

The Peak List and associated documents will be updated. If the peak is a calibration standard, the
User is given the option of recalculating the calibration curve. If a new curve is calculated, all
compounds will be re-quantified.

The Summary Window can be formatted to include the Detection Flags for each peak. These
give information about the start and end points of the peak and can have the following values:

b Peak starts or ends on the baseline.

d Peak starts or ends on a drop line.

M The peak start or end point has been manually assigned.

X The calibration point has been excluded from calibration curve.

The default Chromatogram display range can be controlled by selecting the Quantify Menu Bar
Display, Chromatogram command. For more information about setting the default
Chromatogram display range, see the “Selecting the Chromatogram Display Range for the
Quantify Sample Report” section, on page 10-39.

Page 10-33
Quantify

To Exclude Erroneous Calibration Points

If, once the calibration curves have been created, a calibration point is seen to be erroneous, it can
be removed from the calibration as follows:

1. Select the Quantify Menu Bar Edit, Calibration Curve command. The Calibration Curve
Edit dialog is invoked; this displays a list of the calibration points used to create the
calibration curve. Each point is displayed with Peak List name, standard concentration,
residual error % and a label to indicate whether the point has been included or excluded from
the current calibration curve.

Figure 10.25 The Calibration Curve Edit dialog

2. To exclude a point from the calibration curve, select the calibration point in the list and select
the Exclude button. The label for the point will change from Include to Exclude.

3. To include a point currently not being used in the calibration curve, select the calibration
point in the list and select the Include button. The label for the point will change from
Exclude to Include.

4. Select the OK button to save the changes. The Calibration Curve Edit, Quantify
Compounds? dialog is invoked. Select Yes to quantify compounds, or No to keep the
existing calculated concentrations.

Figure 10.26 The Calibration Curve Edit, Quantify Compounds? dialog

The calibration curve will be re-plotted using only the included calibration points. An
excluded point is denoted by a circle around the point. In the Summary Reports, an excluded
point is denoted by an X in the Detection Flags column.

Page 10-34
 Quantify

To Prevent a Complete Sample from being Used to Create the

Calibration Curve

If, once the calibration curves have been created, all calibration points from a particular standard
sample are seen to be erroneous, the sample can be removed from the calibration as follows:

1. Determine which sample produced the erroneous calibration points.

2. In the Sample List Editor, find the row containing the erroneous sample and set the Sample
Type field to Blank. Alternatively, remove the row from the Sample List.

3. Select the Quantify Application Bar, Process Samples icon; the Quantify Samples dialog is
invoked.

4. Select the Calibrate Standards, Quantify Samples and Print Quantify Report options.
There is no need to select the Integrate Samples option again.

5. Select the OK button; the analysis will start.

To Perform Any of the Quantify Processes

1. Select the Quantify Menu Bar Process, Calculate command. The Quantify Process dialog is
invoked.

Figure 10.27 The Quantify Process dialog

2. Select the required Quantify processes by selecting the relevant check boxes.

Locate Locates peaks for all compounds in the current method.

compounds
Calculate Plots calibration curves for all standards.
calibration
curves
Quantify Calculates concentrations for analyte samples using the current calibration
compounds curves.

Blank subtract When a sample defined as a Blank is encountered, the value is saved and
compounds subtracted from subsequent samples until the next Blank is encountered;
this new value is saved and subtracted from the next set of samples.

3. Select the OK push-button; the selected processes are started.

Page 10-35
Quantify

Printing Quantify Reports

General

To print Quantify Reports, select the Quantify Menu Bar File, Print Report command; the
Quantify Reports dialog is invoked.

Figure 10.28 The Quantify Reports dialog

Reports Frame

Quantify Prints quantification results for each of the Quantify compounds, ordered
summary by by compound.
compound
Quantify Prints quantification results for each of the Quantify compounds, ordered
summary by by sample.
sample
Calibration Prints a calibration curve graph for each Quantify compound.
Curves
Quantify Prints graphs of all the located chromatogram peaks and tables of
samples quantification results. The report is grouped by sample.

Note:
Chromatogram is invoked when producing the report.

Sample Range
Frame
Start…End Enter the Sample numbers for the range to be included in the report.

Page 10-36
 Quantify

Format Invokes the Quantify Report Format dialog; this allows the format of the
Quantify Reports to be changed. See the “Changing the Printed Quantify
Reports Format” section, below, for details.

OK Invokes the Print Report dialog.

Figure 10.29 The Print Report dialog

Printer: Displays the currently selected printer; select the Setup push-button to
change this, see below.

Trace Width Select a trace width from this list box; larger values give larger trace
widths.

Report Font Enter the required value for the printed report font size.
Point Size
Setup Invokes the standard windows Print Setup dialog.

Margins Invokes the Print Margins dialog; this allows the printed report margins
to be set.

OK Prints the reports.

Changing the Printed Quantify Reports Format

Select the Quantify Menu Bar File, Report Format command, or select the Quantify Reports
dialog Format button (see the “Printing Quantify Reports” section, on page 10-36); the Quantify
Report Format dialog is invoked.

Orientation The print orientation for each type of report can be individually specified
Frame by selecting the required option from the list boxes.

Sample Report
Frame
Horizontal Enter the number of horizontal graphs to be displayed in each report.
graphs

Page 10-37
Quantify

Figure 10.30 The Quantify Report Format dialog

Vertical Enter the number of vertical graphs to be displayed in each report.

Graphs
Display Table Prints a summary table of the results, in addition to the graphs.

Display Prints the Internal Standard Chromatogram with the Analyte

Internal Chromatogram. The Internal Standard for a compound is specified in the
Reference Method Editor dialog, Internal Ref field.

Summary by
Sample Report
Frame
Display Totals Displays the breakdown of total compounds for each sample report.

Page Frame

Header: Enter the header text that will appear on each printed page.

Footer: Enter the footer text that will appear on each printed page.

Page numbers Inserts page numbers on each printed page.

Report formats and Quantify Summary formats can be saved and retrieved from the Quantify
Window. This allows the creation of specific summary and report formats to display different
types of data.

Page 10-38
 Quantify

Selecting the Chromatogram Display Range for the

Quantify Sample Report
Select the Quantify Menu Bar Display, Chromatogram command; the Quantify
Chromatogram Display dialog is invoked.

Figure 10.31 The Quantify Chromatogram Display dialog

Show Internal Displays the internal reference with the current peak.
Reference
Add to existing Adds each new chromatogram trace to those already displayed.
chromatograms
Add Adds each new chromatogram trace to the bottom of the previous trace.
chromatograms
top to bottom
Display Range From the list box, select Integration to use the range which was
integrated over, Acquisition to use the range acquired over and Keep
Current to use the current range.

Copying Quantify Summary Reports to the

Clipboard
Quantify allows the equivalent of the Quantify Summary Report to be copied to the Clipboard.
Quantify uses the currently selected Sample List, Method and Peak List files.

Two options are available, the Quantify Summary Report can be ordered either by compound, or
by sample.

To copy the Quantify Summary information to the clipboard, select the Quantify Menu Bar Edit,
Copy Summary By Compound or Copy Summary By Sample command, as appropriate.

Exporting to a LIMS File

General

Quantification results can be written to a text file for use with LIMS systems. This can be
performed automatically by selecting the Quantify Samples dialog, Export Results to LIMS
option, see the “4. Quantify the Data” section, on page 10-22. The results can also be exported
from the Quantify window. Select the Menu Bar File, Export to LIMS File command, select a
file from the invoked browser, or enter the name of a new one and select Save.

Page 10-39
Quantify

The file generated consists of three areas: the Header Section, the Samples Section and the
Calibration section.

The Header Section

The Header Section contains the following four sections. Each shows the full path name of the
file generated by or used to create the report and the date and time that the file was last modified.

• LIMS EXPORT FILE The LIMS file generated.

• SAMPLELIST The Sample List file.

• QUANMETHOD The quantification method file.

• QUANCALIBRATION The quantify curve file.

The Samples Section

The Samples Section contains an entry for each sample in the current Sample List. For each
sample there is one entry for each compound named in the compound box in the Quantify Method.
Each entry has the following fields, separated by a comma.

• The compound number shown in the compound box in the Quantification Method.

• The text name of this compound.

• The scan at which the matching peak was found in the current sample data file.

• The retention time of the matching peak.

• The relative retention time to the referenced peak at which the matching peak was found.

• The area of the matching peak.

• The height of the matching peak.

• The response of the sample for this compound.

• The flags associated with the peak.

• The concentration of compound recorded for this sample.

• The blank subtracted concentration of the compound for this sample.

• The chromatogram trace used to locate peaks for this compound.

• The error between the expected concentration and the calculated concentration for this sample
for a fixed concentration compound.

• The ordinal number of the compound in the quantification method that is used as the
reference peak for this compound.

• The area of the reference peak.

• The height of the reference peak

Page 10-40
 Quantify

• The retention time of the reference peak.

• The modification date of the peak used to quantify this compound for this sample. This refers
to manually modifying the peak, for example by double clicking on the entry in the peak
display in the quantification window.

• The modification time of the peak.

• The modification text (modification comment) of the peak.

• The MassLynx User who altered the peak.

• The mass of the peak.

• The retention time the peak was expected at for this compound.

• The relative retention time the peak was expected at for this compound.

• The user factor associated with this compound.

• The user RF factor associated with this compound.

• Start retention time of the detected peak.

• End retention time of the detected peak.

The Calibration Section

The Calibration Section contains a subsection for each calibration curve calculated for the current
quantification calibration.

Each subsection contains information as displayed on the calibration graphs window. Where a
line entry is inappropriate it is not entered in the report file.

• Correlation coefficient: or Coefficient of Determination:

• Response Factor: or Calibration Curve:

• Response Type:

• Curve Type:, Origin:, and Weighting:

Files Used During Quantify

General

The Quantify program uses four types of files; these are Sample List, Quantify Method, Peak List
and Calibration Curve. It is recommended that Projects are used when performing quantitation, as
this allows the data to be easily stored and accessed.

The Sample List (.SPL) File

Three items in the Sample List are required for quantitation:

Page 10-41
Quantify

File Name Specifies the sample data file name, which will be the same name as the
corresponding Peak List file.

Sample Type Specifies the type of sample. This should be set to Standard if the sample
is to be used to form a calibration curve, Analyte if the concentration of
the compounds within the samples is to be calculated, QC if it is a Quality
Control sample, or Blank if the sample doesn’t contain any analyte
compounds.

Concentration Only required if the sample is a Standard and is optional for QC samples.
Specifies the known concentrations of the compounds within the standard.
This does not apply to compounds whose concentration has been specified
as being constant (Fixed), within all samples.

The Sample List files are normally stored in the \SAMPLEDB directory.

The Quantify Method (.MDB) File

The Quantify Method contains an entry for each of the compounds being analyzed, determining
how the data is to be processed. The same method is applied to all the samples in the analysis.
For more information, see the “2. Create a Quantify Method” section, on page 10-11. The
Quantify Method files are normally stored in the \METHDB directory.

Peak Lists (.PDB) File

A Peak List contains peaks that were detected when integrating chromatograms. Further
information gathered as a result of running Quantify, such as compound name and concentration,
are also saved in the Peak List.

Peak Lists are produced as a result of running the MassLynx automated Quantify software, or by
the Chromatogram service. One Peak List should be formed for each of the samples in the
analysis, the Peak List will have the same name as the sample from which it was formed.

For more information on examining, modifying and creating Peak Lists see Chapter 6,
“Chromatogram”.

The Peak List files are normally stored in the \PEAKDB directory.

Calibration Curves (.CDB) File

Stores the Quantify Calibration Curves which are produced for each of the compounds within the
method. The Calibration Curve file has the same name as the method used to create it. The
Calibration files are normally stored in the \CURVEDB directory.

Page 10-42
 Library

Chapter 11 Library

Page 11-1
Library

Contents
Introduction ................................................................................................................................. 11-5
The Library Tool Bar .................................................................................................................. 11-6
Library Searching ........................................................................................................................ 11-6
General............................................................................................................................. 11-6
The Presearch................................................................................................................... 11-6
The Mainsearch................................................................................................................ 11-7
An Overview of Library Searching .................................................................................. 11-7
Selecting Which Libraries to Search ................................................................................ 11-8
Selecting a New Search Spectrum.................................................................................. 11-10
Changing the Library Search Parameters ....................................................................... 11-11
Library Search Filters..................................................................................................... 11-13
Starting a Library Search ............................................................................................... 11-15
Automatic Library Search .............................................................................................. 11-15
Library Search Results .............................................................................................................. 11-16
General........................................................................................................................... 11-16
Manipulating the Library Display .................................................................................. 11-17
The Hit List Window................................................................................................................. 11-18
General........................................................................................................................... 11-18
Formatting the Hit List................................................................................................... 11-19
The Hits Window ...................................................................................................................... 11-21
General........................................................................................................................... 11-21
Manipulating the Hits Window Display......................................................................... 11-21
The Delta Window .................................................................................................................... 11-22
The Structure Window .............................................................................................................. 11-23
Printing the Results of a Library Search.................................................................................... 11-23
Copying to and from the Windows Clipboard........................................................................... 11-24
To Copy a Picture to the Clipboard................................................................................ 11-24
To Copy the Current Hit List to the Clipboard............................................................... 11-24
To Retrieve Data from the Clipboard............................................................................. 11-24
Refining the Search Spectrum ................................................................................................... 11-24
General........................................................................................................................... 11-24
To Refine the Search Spectrum...................................................................................... 11-25
Auto Refine .................................................................................................................... 11-25
Library Compare Process .......................................................................................................... 11-25
General........................................................................................................................... 11-25
To Compare the Search Spectrum with a Particular Library Entry................................ 11-25
Library Subtract Process ........................................................................................................... 11-26
General........................................................................................................................... 11-26
To Subtract a Particular Hit from the Search Spectrum ................................................. 11-26
User Libraries ............................................................................................................................ 11-26
General........................................................................................................................... 11-26
To Create a New User Library ....................................................................................... 11-26
To Append a Spectrum to a User Library ...................................................................... 11-28
To Add Spectra from a Library to an Existing User Library.......................................... 11-28
To Create a Spectrum and Add it to an Existing User Library....................................... 11-29
Adding Textual Data to Library Entries......................................................................... 11-29
Indexing a User Library ................................................................................................. 11-31
Deleting Library Entries................................................................................................. 11-32
The Library Locator .................................................................................................................. 11-32
General........................................................................................................................... 11-32
To Select a Particular Entry for Display ........................................................................ 11-33
To Locate Entries with Filters........................................................................................ 11-33
To Set the Locate Filters ................................................................................................ 11-33

Page 11-2
 Library

Illustrations
Figure 11.1 Typical Library Window......................................................................................... 11-5
Figure 11.2 The Library Search List dialog ............................................................................... 11-8
Figure 11.3 The Add Library dialog .......................................................................................... 11-9
Figure 11.4 The Library Data Browser dialog ......................................................................... 11-10
Figure 11.5 The Scan Select dialog.......................................................................................... 11-11
Figure 11.6 The Library Search Parameters dialog.................................................................. 11-11
Figure 11.7 The Filters dialog .................................................................................................. 11-13
Figure 11.8 The Edit Peak List dialog...................................................................................... 11-16
Figure 11.9 The Library Display View dialog ......................................................................... 11-17
Figure 11.10 Typical Hit List Window .................................................................................... 11-18
Figure 11.11 The Format DB List dialog ................................................................................. 11-19
Figure 11.12 The List Field Justification dialog ...................................................................... 11-20
Figure 11.13 Typical Hits Window.......................................................................................... 11-21
Figure 11.14 The Display Data dialog ..................................................................................... 11-22
Figure 11.15 Typical Delta Window........................................................................................ 11-22
Figure 11.16 Typical Structure Window .................................................................................. 11-23
Figure 11.17 The Refine Spectrum dialog ............................................................................... 11-24
Figure 11.18 The Library Compare dialog............................................................................... 11-25
Figure 11.19 The Subtract Hit dialog....................................................................................... 11-26
Figure 11.20 The Append Spectrum dialog ............................................................................. 11-27
Figure 11.21 The Append File Select dialog............................................................................ 11-27
Figure 11.22 Typical Append File Select prompt .................................................................... 11-27
Figure 11.23 Typical Append Spectrum prompt...................................................................... 11-27
Figure 11.24 The Display Library Spectrum dialog................................................................. 11-28
Figure 11.25 The Library Edit dialog....................................................................................... 11-29
Figure 11.26 The Library Reindex dialog ................................................................................ 11-31
Figure 11.27 The Library Locator dialog ................................................................................. 11-32

Page 11-3
Library

Page 11-4
 Library

Introduction
The MassLynx Library service is used to identify a search (unknown) spectrum by comparing it
with a library of known spectra. The result of a Library search is a list of library compounds, or
“hits”, whose spectra give the best match with the unknown spectrum.

The Library Window uses multiple windows to present the search results in several formats.

• The Hit List Window gives a textual listing of the best hits. The Hit List Window display
can be formatted to display information about each hit including compound name, fit values,
formula, molecular weight, etc. See the “The Hit List Window” section, on page 11-18.

• The Hits Window shows the search spectrum followed by the spectra of the best hits. See the
“The Hits Window” section, on page 11-21.

• The Delta Window shows the difference between the search spectrum and the spectrum of a
particular hit. See the “The Delta Window” section, on page 11-22.

• The Structure Window shows the chemical structure of the currently selected hit. See the
“The Structure Window” section, on page 11-23.

Library also allows the User to create their own User Libraries containing spectra from raw data
files via the Spectrum Window. See the “User Libraries” section, on page 11-26.

Libraries can be edited via the Library Edit dialog; see the “Adding Textual Data to Library
Entries” section, on page 11-29.

The Library Locator dialog can be used to examine a library and search for library entries that
meet various criteria; see the “The Library Locator” section, on page 11-32.

To access the Library, select the MassLynx Tools Shortcut Bar, Search Library icon.

Figure 11.1 Typical Library Window

Page 11-5
Library

The Library Tool Bar

The Library Tool Bar is displayed at the top of the Library Window.

Tool Menu Command Function

Bar
Button
File, Open Opens a data file.

File, Print Prints the current window in portrait format.

File, Print Prints the current window in landscape format.

Edit, Copy Bitmap Copies a bitmap of the current window to the clipboard.

Edit, Copy Hit List Copies the current hit list to the clipboard.

Process, Refine Refines the current search spectrum.

Process, Search Searches the current search spectrum against the current
library.

Window, Tile Arranges the windows in a tiled view.

Window, Cascade Arranges the windows in a cascaded view.

Window, Stack Arranges the windows in a stacked view.

Display, Spectrum Selects a new scan number as the search spectrum.

Press once to restore the previous display range; press

again to use the default display range.

Library Searching
General

The Library Search process has two parts - the Presearch and the Mainsearch. The Presearch is a
quicker search that is designed to select a number of likely candidates from the library. These
candidates are then passed through to the Mainsearch where they undergo a more rigorous and
lengthy comparison. The results of the Mainsearch are then displayed in the Hit List, Hits, Delta
and Structure windows.

The Presearch

The Library Presearch file contains a spectrum, which has been reduced to the eight most intense
mass-weighted peaks, for each library entry. The search spectrum is likewise reduced to its eight
most intense mass-weighted peaks and then compared to the Library Presearch file spectra. The
most likely candidates are those compounds with spectra that have the greatest number of
matching peaks with the unknown compound. A list of the most likely candidates is passed to the

Page 11-6
 Library

Mainsearch process. The User can control how many candidates are passed to the Mainsearch by
altering the Library Search Parameters dialog, Match by parameter.

The Mainsearch

For the Mainsearch, the search spectrum is again reduced, this time to a number of peaks, which is
selected by the User via the Library Search Parameters dialog, Sig. Peaks parameter.

The search spectrum is compared to each of the possible candidates from the Library and the
results of this comparison are presented in the Hits, Hit List, Delta and Structure windows. The
hits are ranked in order of best fit to the search spectrum.

Various filters can be applied to the Mainsearch process to make it more specific. These filters
contain requirements, such as elemental formula and molecular weight, which must be met before
the Library entry can be included in the list of hits.

Two types of fit values are computed for each hit. These are Forward and Reverse fit. The
maximum obtainable fit value is 1000, which represents a perfect match between the search
spectrum and the Library entry.

The Forward fit value shows how likely it is that the search spectrum is a pure sample of the
Library entry. Any peaks that are present in the search spectrum, but not present in the Library
spectrum, will decrease the Forward fit value. Likewise, any peaks that are present in the Library
spectrum, but not present in the search spectrum, will decrease the Forward fit value.

The Reverse fit value shows how likely it is that the search spectrum contains the Library entry; in
this case the search spectrum may be a mixture of compounds. Any peaks that are present in the
Library spectrum, but not present in the search spectrum, will decrease the Reverse fit value.
However, peaks that are present in the search spectrum, but not present in the Library spectrum,
will have no effect on the reverse fit value.

An Overview of Library Searching

This section lists the steps involved in a Library search. Each step is covered in more detail within
its own section later in this User’s Guide.

1. Select the Library or Libraries to use for the search using the Library Menu Bar File, Search
List command.

2. Select the search spectrum. The search spectrum can either be displayed in the Spectrum
window or in the Library window. A new data file can be selected within Library by
selecting the Tool Bar button or using the Menu Bar File, Open command to invoke the
Library Data Browser dialog. A new scan can be selected from the current data file by
using the Tool Bar button or using the Menu Bar Display, Spectrum command.

3. Edit the Library search parameters using the Menu Bar Edit, Parameters command.

4. Apply any search filters using the Menu Bar Edit, Filters command.

5. Initiate the Library search. The Library search can be started either from Library or Spectrum
using the Tool Bar button, or by using the Library Menu Bar Process, Search command.

6. Adjust the Library display using the Menu Bar Display, View command. Format the Hit List
document using the Menu Bar Edit, Format List command.

Page 11-7
Library

7. Print the results of the Library search by selecting the Tool Bar or button, or using
the Menu Bar File, Print command.

Note:
All the above settings will be retained for future searches and only need to be edited when they
require changing.

Selecting Which Libraries to Search

General

The Library program will search one or more Libraries specified in its Library Search List.
These can be standard Libraries such as the NIST Library, or User Libraries (created by the User).
The current search list is displayed in the Library Search List dialog, which is invoked by the
Menu Bar File, Search List command.

Figure 11.2 The Library Search List dialog

Search Method
Frame
MassLynx Selects the MassLynx Library search procedure.

NIST Selects the NIST Library search procedure.

Library Search Lists all of the libraries that are associated with the current search process.
List
OK When selected, the search procedure will default to that selected in the
Search Method Frame, with the libraries to be searched being those in the
corresponding Library Search List.

Note:
This button is disabled when the search list dialog is invoked; it is enabled
when a change has been made to the dialog.

Add Invokes the Add Library dialog, see the “To Add a New Library to the
Current Search List” section, on page 11-9.

Page 11-8
 Library

Remove Removes the currently selected Library from the Library Search List, see
the “To Remove a Library from the Current Search List” section, on
page 11-9.

To Add a New Library to the Current Search List

1. Select the Menu Bar File, Search List command; the Library Search List dialog is invoked.

2. Select the required Search Method; MassLynx or NIST.

3. Select the Add button:

For a MassLynx search, the Add Library dialog is invoked.

Figure 11.3 The Add Library dialog

a. Select the required file.

b. Select the Open push-button; the Add Library dialog is closed and the library is added
to the Library Search List dialog Library Search List.

For a NIST search, the standard Windows Browse for Folder dialog is invoked.

a. Navigate to the folder containing the required Library.

b. Select the Open push-button; the Browse for Folder dialog is closed and the library is
added to the Library Search List dialog Library Search List.

4. Select the OK push-button; the Library Search List dialog is closed.

To Remove a Library from the Current Search List

1. Select the Menu Bar File, Search List command; the Library Search List dialog is invoked.

2. Select the Library to be removed.

3. Select the Remove button. The Library is deleted from the Library Search List list box.

4. Select the OK push-button; the Library Search List dialog is closed.

Page 11-9
Library

Selecting a New Search Spectrum

To Select a Spectrum from a Different Data File

1. Select the Tool Bar button, or select the Menu Bar File, Open command; the Library
Data Browser dialog is invoked.

Figure 11.4 The Library Data Browser dialog

2. Select the new data file in the File Name: list box. A processed spectrum, which is the result
of Combine or Refine processes, can be selected using the History button. For more detailed
information about using the Data Browser, see Chapter 3, “The MassLynx Window and
Related Information”.

3. Select OK to exit the Library Data Browser.

4. The Library Hits window will be updated to show scan 1 of the new data file; this will
become the current search spectrum. See the “The Hits Window” section, on page 11-21.

Page 11-10
 Library

To Select a New Scan from the Current Data File

1. Select the Tool Bar button, or select the Menu Bar Display, Spectrum command; the
Scan Select dialog is invoked.

Figure 11.5 The Scan Select dialog

2. Type the required scan number, or retention time in the Entry box.

3. Select the Update button to update the spectrum displayed in the Library Hits window.

4. Select the OK push-button.

To Select a New Search Spectrum from the Spectrum Window

If the Library search is initiated from the Spectrum Window, the spectrum currently displayed in
that Window will be used as the search spectrum. New spectra can be selected in the Spectrum
Window using the Menu Bar File, Open command, the Display, Spectrum, At command, or the
Tool Bar button. For more detailed information, see Chapter 7 “Spectrum”.

Changing the Library Search Parameters

The Library Search parameters control how many Library entries are passed from the Presearch to
the Mainsearch, exactly which entries are used and how the results are reported.

The Library Search parameters are accessed by selecting the Menu Bar Edit, Parameters
command, this invokes the Library Search Parameters dialog.

Figure 11.6 The Library Search Parameters dialog

Page 11-11
Library

Match By Frame The Match By parameter determines how many candidates from the
Library will be passed from the Presearch to the Mainsearch.

Level: When selected, all Library entries having the number of matching peaks,
or above, specified in the adjacent text box, will be passed to the
MainSearch process. The higher the number entered, the fewer entries
will be passed to the Mainsearch.

Note:
The text box will only accept integer values between 0 and 8.

Viables: When selected, the minimum number of entries that will be passed from
the Presearch to the Mainsearch is specified in the adjacent text box.
Library first takes all entries that have eight matching peaks. If the
number of entries is less than the Viables value, Library takes all entries
that have seven matching peaks; it adds these to the previous entries and
compares the new total to the Viables value. This process is repeated until
the number of entries is greater than, or equal to, the Viables value. In
practice, the number of entries passed to the Mainsearch is often much
larger than the Viables value.

Sig. Peaks Frame

Number: The value of this parameter determines how many peaks are to be used to
create the reduced spectrum following the possible exclusion of any
masses. For example, if the value is 20, then after mass weighting the
peaks the top twenty will become the reduced spectrum.

Note:
The text box will only accept integer values between 0 and 200.

Ranking Frame The Ranking parameter determines whether hits will be listed in order of
Forward: or Reverse: fit.

Forward: Shows how likely it is that the search spectrum is a pure sample of the
Library entry. Any peaks, which are present in the search spectrum but
not present in the Library spectrum, will decrease the Forward fit value.
Likewise, any peaks that are present in the Library spectrum, but not
present in the search spectrum will also decrease the Forward fit value.

Reverse: The Reverse fit value shows how likely it is that the search spectrum
contains the Library entry, in this case the search spectrum may be a
mixture of compounds. Any peaks, which are present in the Library
spectrum but not present in the search spectrum, will decrease the Reverse
fit value. However, peaks, which are present in the search spectrum but
not present in the Library spectrum, will have no effect on the Reverse fit
value.

Page 11-12
 Library

Exclude Masses The Exclude Masses parameter allows the exclusion of up to four
Frame particular masses in the search spectrum from the Mainsearch (Mass 1: to
Mass 4:; enter the required masses in the adjacent text boxes). These
excluded peaks will not be compared to Library entries. This can be
useful, for example, to exclude a contaminating ion, which cannot be
removed from the spectrum by any physical or chemical means.

The All Below parameter allows all masses below a certain value to be
excluded.

Library Search Filters

The Library search filters are used to specify certain criteria that a Library entry must meet before
it will appear in the Hit list. If a compound’s molecular weight and elemental formula is known,
these filters can be used to make the search more specific. For example, if the search compound is
known to contain at least one chlorine atom, this can be specified in the search filters.
Alternatively, if it is known that its molecular weight must lie within a certain range, this can be
specified.

Select the Menu Bar Edit, Filters command to invoke the Filters dialog.

Figure 11.7 The Filters dialog

Fit Frame The Fit parameters allow the User to specify a Minimum Forward and/or a
Minimum Reverse fit value, which a Library entry must have before it will
appear in the Hit list.

Min Forward The Minimum Forward filter, enter a value between 0 and 1000 in the
adjacent text box.

Min Reverse The Minimum Reverse filter, enter a value between 0 and 1000 in the
adjacent text box.

Page 11-13
Library

Mol Wt Frame Specifies a range within which the molecular weight of the Library entry
must fall before it will be included in the Hit list. To activate the filter,
select the Active option and enter the minimum and maximum molecular
weights in the Min and Max text boxes. To specify a particular molecular
weight, make the Min and Max values equal.

Elements Frame The Elements filter specifies minimum and maximum numbers of
particular elements that must be present in the Library entry’s molecular
formula before it will appear in the Hit list.

Elemental formulae are entered in ‘standard’ format as an element symbol,

followed immediately by its count if greater than one, and then
immediately by another symbol, as relevant. To make the filter active,
select the Active box.

For example, consider Elements set to C6H20NClBr2. Symbols must be

entered in correct upper and lower case format. Note also that, “Cl” does
not need a “1” after it and that there are no spaces. The specific order of
elements is irrelevant.

If an element appears in the Min control then the library entry must
contain the specified number of atoms, e.g. Cl2, but can contain any
number above this, e.g. Cl3, Cl4, etc.

If an element appears in the Max control then the library entry must not
contain more than the specified number of atoms, e.g. Cl2, but can contain
any number below this, e.g. no Cl atoms, Cl or Cl2.

If an element appears in both the Min and Max controls, then the number
of atoms must lie between the two values specified. If the values are the
same then the library entry must contain exactly this number of atoms.

To match a specific formula, enter the formula in both the Min and Max
controls and do not select Include Other Elements. If Include Other
Elements is selected then other elements may be present in the Library
entry.

Flags Frame Note:

This Frame is only relevant to User Libraries; it is only available if there
is a User Library in the Search List.

The Flags parameter specifies a range of values within which a Library

entry’s Flags must lie before it will appear in the Hits list. These Flags are
strings of one or more characters that have been entered in the User
Library.

User Flags Enter the required flag text, eight characters maximum.

Page 11-14
 Library

Apply exact When selected, the User Flags of a library entry have to match exactly
with those given; including lower/upper case, in the specified order.

When not selected, the Library entry must contain the characters specified
in the User Flags control, however, these characters can appear in any
order in the matching library entry.

For example, if the User Flags control is set to Bv, any of the following
will pass a non-exact search; BpKv, vKpB or KBvp.

User Values Note:

Frame
This Frame is only relevant to User Libraries; it is only available if there
is a User Library in the Search List.

The User Values parameter specifies a range of values within which a

Library entry’s User Values must lie before it will appear in the Hits list.
These are numeric values that have been entered in the User Library.

Select the No. 1 and No. 2 value controls as required. A maximum and
minimum value for each User Value can be entered in the Min and Max
controls. To specify a particular User Value, make the Min and Max
values equal.

Starting a Library Search

A Library search can be started from either Library or Spectrum.

To start a library search from Library, select the Tool Bar button, or select the Menu Bar
Process, Search command.

To initiate a Library search from Spectrum, select the Tool Bar button, or select the Menu Bar
Tools, Library Search command.

Automatic Library Search

General

The library search module used for identifying spectra by matching them with a standard library
(e.g. NIST) currently works on a single spectrum. A facility to automatically search for a number
of spectra from a data set has been added.

To Use Automatic Library Search

1. In the Chromatogram window, integrate the chromatogram of interest.

2. In the Chromatogram window, select the Menu Bar Edit, Peak List Write command; the
Edit Peak List dialog is invoked.

3. Select the required Peak Top and select the Append button, repeat this for each peak
required, or select the Append All button to append all peaks.

4. Select the Exit push-button; the Edit Peak List dialog is closed.

Page 11-15
Library

Figure 11.8 The Edit Peak List dialog

5. In the Library Window, select the Menu Bar Process, Auto Refine command.

6. Select the Menu Bar Process, Search Peak List command.

7. The Library search process performs a search for each peak in the list and displays the Print
dialog. Select All Windows to print results for all windows, or Current Window to print
results for the current window, and select the OK button to print.

Library Search Results

General

The result of a Library search is a list of library compounds, or “hits”, whose spectra give the best
match with the unknown spectrum.

The results are displayed in four windows.

The Hit List Window gives a textual listing of the best hits. The Hit List window display can be
formatted to display a variety of information about each hit including compound name, fit values,
formula, molecular weight etc. See the “The Hit List Window” section, on page 11-18.

The Hits Window shows the unknown spectrum followed by the spectra of the best hits; see the
“The Hits Window” section, on page 11-21.

The Delta Window shows the difference between the unknown spectrum and the spectrum of the
current hit, see the “The Delta Window” section, on page 11-22.

The Structure Window shows the chemical structure of the currently selected hit, see the “The
Structure Window” section, on page 11-23.

Page 11-16
 Library

Manipulating the Library Display

The appearance of the Library display can be altered by selecting the Library Display, View
command; the Library Display View dialog is invoked.

Figure 11.9 The Library Display View dialog

To display any of the windows, select the Visible option for that window.

The Peak Label Threshold controls alter peak labeling in the Hits and Delta windows. The
number of decimal places to which peaks are labeled (from 0 to 4) is entered in the Decimal
Places control. A threshold can be set for labeling peaks with mass. Selecting None results in no
mass labels for any peaks. A relative intensity threshold for peak labels can be set by selecting the
% Full Scale control and entering a % value. An absolute intensity threshold for peak labels can
be set by selecting the Intensity control and entering an intensity value.

For the Hits window, the number of Hits displayed with the search spectrum can be selected. This
is selected in the No. Hits control in the range 1 to 4.

It is also possible to edit the header displayed at the top of each window by selecting the Header
button. This will invoke the Header Editor. For more information about using the Header
Editor, see Chapter 3, “The MassLynx Window and Related Information”.

The different Library documents can be arranged within the Library window using the commands
in the Window menu.

Page 11-17
Library

The Hit List Window

General

Figure 11.10 Typical Hit List Window

The Hit List Window gives a textual listing of the best twenty hits resulting from the Library
search. These hits can be listed in order of either Reverse or Forward fit depending on which
order was selected for the Library Search Parameters dialog Ranking parameter. The Hit List
document can be formatted to include the following information about each hit:

• Hit number.

• Compound name.

• Forward fit value.

• Reverse fit value.

• Chemical formula.

• Molecular weight.

• Library entry number.

• Library.

• Chemical Abstracts Sequence (CAS) number.

Click on a hit in the Hit List Window to make it current. The current hit will be also shown in
Delta and Structure Windows, see the “The Delta Window” section, on page 11-22, and the “The
Structure Window” section, on page 11-23.

Page 11-18
 Library

Formatting the Hit List

Introduction

Select the Menu Bar Edit, Format List command; the Format DB List dialog is invoked.

Figure 11.11 The Format DB List dialog

The fields currently used in the Hit List document are shown in the Format list. Fields that can be
added to the Hit List document are shown in the Fields list.

To Append New Fields to the Hit List

1. Select the required field in the Fields list box.

2. Select the Append button.

3. To view the result of this change without exiting the dialog, select the Update button.

4. Repeat steps 1 to 3, as required.

5. Select the OK button to save the changes and exit.

To Insert New Fields in the Hit List

1. Highlight the field to be inserted in the Fields list box.

2. In the Format list box, select the field before which the new field is to be inserted.

3. Select the Insert button.

4. To view the result of this change without exiting the dialog, select the Update button.

5. Repeat steps 1 to 4, as required.

6. Select the OK button to save the changes and exit.

Page 11-19
Library

To Remove a Field from the Hit List Document

1. Highlight the field to be removed in the Format list box.

2. Select the Remove button. To remove all the fields in the Hit List document select the
Remove All button.

3. To view the result of this change without exiting the dialog, select the Update button.

4. Repeat steps 1 to 3, as required.

5. Select the OK button to save the changes and exit.

To Alter the Justification of a Field in the Hit List Document

1. Highlight the appropriate field in either the Fields or Format list boxes.

2. Select the Justification button; the List Field Justification dialog is invoked.

Figure 11.12 The List Field Justification dialog

3. Select the required justification: Left, Centre or Right.

4. Select the required Field Width, Significant Figures (SF) or Decimal Places (DP).

5. Repeat steps 1 to 4, as required.

6. Select the OK button to save the changes and exit.

Page 11-20
 Library

The Hits Window

General

Figure 11.13 Typical Hits Window

The Hits Window displays the search spectrum with up to four of the hits spectra.

The header above each hit spectrum shows the hit number, fit value, the Library and Library entry
number and the compound name.

The mass axis can be zoomed to expand a region of particular interest, these changes will also be
reflected in the Delta document.

Manipulating the Hits Window Display

To Determine which Hits are Displayed

The first hit displayed is always the current hit, which is the hit highlighted in the Hit List
Window. The Hits Window will display up to four of the next best hits. The number of hits
displayed is altered by selecting the Menu Bar Display, View command and changing the No.
Hits parameter value.

Altering the Range of the Mass Axis with the Mouse

Press the mouse button at one end of the region of interest, and without releasing the button, drag
the cursor horizontally to the other end. As the cursor is dragged, a “rubber band” is stretched out
to indicate the range selected; do not go beyond the bounds of the axis. When the mouse button is
released, the selected range will be re-displayed to fill the current window.

This operation can be repeated as often as required.

Page 11-21
Library

Restoring the Display from the Tool Bar

Pressing the Tool Bar button once restores the display to its previous state. Pressing it a
second time restores the display to the default range.

To Alter the Range of the Mass Axis using the Menu Bar

1. Select the Menu Bar Display, Range, From command; the Display Data dialog is invoked.

Figure 11.14 The Display Data dialog

2. Enter new From and To values for the mass axis.

3. Select the OK button.

Restoring the Display using the Menu Bar

Select the Menu Bar Display, Range Default command.

The Delta Window

Figure 11.15 Typical Delta Window

The Delta Window shows the difference between the search spectrum and the currently selected
hit. Positive peaks are those which are more intense in the search spectrum than the hit spectrum.
Negative peaks are those which are more intense in the hit spectrum than the search spectrum.

Page 11-22
 Library

The 100% annotation point of the intensity axis refers to the base peak intensity of the spectra
prior to subtraction.

The mass axis of the Delta document is always the same as the Hits document and cannot be
changed independently.

The Structure Window

Figure 11.16 Typical Structure Window

The Structure Window shows a graphical representation of the chemical structure of the currently
selected hit.

The structural pictures are derived from structure data supplied by the National Institute for
Standards (NIST) and are their copyright. Not all NIST Library entries have associated structures.
If the currently selected hit has no associated structure, a message “No structure found” will
appear in the Structure window.

If the Structure Window is blank, it may be because it is too small to contain the structure, try
maximizing the window as a quick check.

Structures are associated to Library entries by their CAS number. If a User Library is created and
the correct CAS numbers entered, the structures for the entries can be viewed.

Printing the Results of a Library Search

The currently selected Library window can be printed in portrait format by selecting the Tool Bar
button, or in landscape format by selecting the Tool Bar button.

The results of a Library search can also be printed by selecting the Library Menu Bar File, Print
command.

Page 11-23
Library

Copying to and from the Windows Clipboard

To Copy a Picture to the Clipboard

1. Select the required window and alter the display as required.

2. Select the Tool Bar button, or select the Menu Bar Edit, Copy Bitmap command.

The displayed picture will now be transferred as a bitmap to the Windows clipboard, and can be
pasted into any Windows compatible software.

To Copy the Current Hit List to the Clipboard

Select the Tool Bar button to copy the current hit list to the clipboard.

To Retrieve Data from the Clipboard

Many Windows applications have an Edit, Paste, or similar command to read data in from the
clipboard. Consult the application’s manual, or help text for more information.

MassLynx Spectrum and Chromatogram services are able to read bitmaps via their Edit Paste
commands.

Refining the Search Spectrum

General

The Refine process operates on centroid-mode data only. Its purpose is to identify just those
masses that contribute to a specific peak in the TIC. In this way, it removes small peaks that are
due to background noise and can improve the results of Library searching. The Refine Spectrum
dialog is invoked by the Menu Bar Process, Refine command.

Figure 11.17 The Refine Spectrum dialog

The User defines two parameters for the Refine process; Window size and Noise threshold.

The refine algorithm proceeds by generating the summed mass chromatogram over a range of
1 Da centered on each integer mass in turn. It examines these chromatograms for a number of
scans equal to the Window size around the Peak top scan. If there is a peak present in this range
whose topmost point is within one scan of the Peak top scan and more intense than the Noise
threshold value, this mass will appear in the refined spectrum.

Page 11-24
 Library

To Refine the Search Spectrum

1. Select the Menu Bar Process, Refine command; the Refine Spectrum dialog is invoked.

2. Enter a value for Window size; this is the half width, in scans, at the baseline of the TIC peak
of interest.

3. Enter a value for Noise threshold. For the first run, set Noise threshold to zero to show all
the peaks.

4. Select the OK button.

5. If the noise level in the refined spectrum is unacceptable, repeat the refine operation with a
higher Noise threshold setting. Values in the range 0 to 10 are recommended.

The current spectrum may also be refined, using the current refine parameters, by selecting the
Tool Bar button.

Auto Refine

To automatically use the refine parameters in all searches, select the Menu Bar Process, Auto
Refine command. A tick mark appears next to the item if it is selected, to turn this option off
select it from the menu again.

Library Compare Process

General

The Compare process allows comparison of the search spectrum with a particular Library entry.
This can be useful if the User has an idea what the compound is, or what type of compound it is,
particularly if the compound in question does not appear in the top twenty hit list.

To Compare the Search Spectrum with a Particular Library Entry

1. Select the Menu Bar Process, Compare command; the Compare dialog is invoked.

Figure 11.18 The Library Compare dialog

2. Enter the Entry Number of the Library entry with which the search spectrum is to be
compared. If required, access a different Library by selecting the File button.

3. Select the OK button.

The Library display will be updated to show the results of the comparison in the Hit List, Hits,
Delta and Structure windows if they are currently displayed. The format of the display is the
same as for a normal search except, of course, there is only one hit.

Page 11-25
Library

Library Subtract Process

General

The Subtract process allows the spectrum of a particular hit to be subtracted from the search
spectrum. The resulting subtracted spectrum becomes the new search spectrum and the Library
search can be repeated.

This can be useful if it is suspected that the search spectrum is a mixture of more than one
compound. This would be indicated by a high reverse fit value and a low forward fit value. If
the spectrum of one of the hits is subtracted from the search spectrum and the Library search
repeated the other component of the mixture should now appear high on the hit list. For mixtures
of more than two compounds this process can be utilized to identify them one at a time.

To Subtract a Particular Hit from the Search Spectrum

1. Select the Menu Bar Process, Subtract Hit command; the Subtract Hit dialog is invoked.

Figure 11.19 The Subtract Hit dialog

2. Enter the number of the hit to be subtracted from the search spectrum.

3. Select the OK button. The subtracted spectrum will become the new search spectrum.

User Libraries
General

As well as the standard NIST library supplied, the User can create their own User Libraries
containing their own spectra. These spectra can come from raw data files, from existing libraries,
or can be created by the User and imported using the DataBridge file conversion program; see
Chapter 13, “DataBridge”.

To Create a New User Library

1. Run the Spectrum program, see Chapter 7, “Spectrum”.

2. Select the first spectrum that is to be included in the library.

3. Select the Spectrum Menu Bar Edit, Library, Append command; the Append Spectrum
dialog is invoked.

Page 11-26
 Library

Figure 11.20 The Append Spectrum dialog

4. Select the File button, the Append File Select dialog is invoked.

Figure 11.21 The Append File Select dialog

5. Enter the name for the new Library in the File name: text box.

6. Select the OK button; a prompt appears.

Figure 11.22 Typical Append File Select prompt

7. Select the YES button to create the new Library; a prompt appears.

Figure 11.23 Typical Append Spectrum prompt

8. Select the OK button to add the first spectrum to the Library.

Page 11-27
Library

9. Append further spectra, as required, to the Library as described in the “To Append a Spectrum
to a User Library” section, below.

10. Add textual data for each Library entry, as required, see the “Adding Textual Data to Library
Entries” section, on page 11-29.

11. Use the Library Menu Bar Process, Index Library command to create the Presearch file for
the Library, see the “Indexing a User Library” section, on page 11-31.

To Append a Spectrum to a User Library

1. Run the Spectrum program, see Chapter 7, “Spectrum”.

2. Select the spectrum that is to be appended to the Library.

3. Select the Spectrum Menu Bar Edit, Library, Append command; the Append Spectrum
dialog is invoked.

4. Select the File button, the Append File Select dialog is invoked.

5. Select the Library to which the spectrum is to be appended.

6. Select the Open button, the Append File Select dialog is closed.

7. Select the OK button to append the spectrum to the Library.

To Add Spectra from a Library to an Existing User Library

1. Run the Spectrum program, see Chapter 7, “Spectrum”.

2. Select the Spectrum Menu Bar Edit, Library, Get Spectrum command; the Display Library
Spectrum dialog is invoked.

Figure 11.24 The Display Library Spectrum dialog

3. Enter the Library Entry: number to be displayed.

4. Select the OK button.

5. Append the spectrum to the Library, as described in the “To Append a Spectrum to a User
Library” section, above.

Page 11-28
 Library

To Create a Spectrum and Add it to an Existing User Library

A spectrum can be created as a text file and imported into MassLynx, using DataBridge, then
appended to a Library.

1. Create the spectrum as a text file containing a list of mass intensity pairs. Any plain text
editor such as Windows Notepad can be used to create the file.

2. Use the DataBridge program to convert the file from ASCII to MassLynx format. See
Chapter 13, “DataBridge” for more details.

3. Append the spectrum to the Library, as described in the “To Append a Spectrum to a User
Library” section, on page 11-28.

Adding Textual Data to Library Entries

General

Select the Menu Bar Edit, Library command; the Library Edit dialog is invoked.

Figure 11.25 The Library Edit dialog

Once a spectrum has been appended to a User Library, it will need editing to add textual data, such
as Compound Name, Text, CAS Number, Formula and Molecular Weight.

Two numerical User Values and User Flags may be added for the entry. These can be used to
hold information about the compound. These fields can then be used as Filters in Library
searches.

Name The compound name for the entry; any text, up to a maximum of
128 characters, can be entered.

Page 11-29
Library

CAS The Chemical Abstracts Sequence (CAS) number for the compound. The
CAS number is used to link Library entries to their chemical structures in
the Structures Library. The CAS number has the format “x-yy-z”, where:

x: Is a string of numbers; e.g., “12398” or “6”;

yy: Is a two-digit number string; e.g., “23” or “07”;

z: Is a one-digit number string; e.g., “7” or “0”.

Formula The elemental formula for the compound.

Elemental formulae are entered in “standard” format as an element

symbol, followed immediately by its count if greater than one, and then
immediately by another symbol, as relevant.

For example, consider Formula set to “C6H20NClBr2”. Symbols should

be entered in correct upper and lower case format. “Cl” does not need a
“1” after it and that there are no spaces. The specific order of elements is
irrelevant.

Mol Wt The molecular weight of the compound and should be entered as an

integer, based on nominal masses for elements; for example, H is 1 and Cl
is 35.

Note:
Formula and Mol Wt are compared within an entry; a warning will be
displayed if there is a discrepancy.

Text Text can be entered up to a maximum of thirty characters.

Value 1 and Any number [positive or negative, integer (no decimal point) or decimal
Value 2 point values] can be entered.

These values can be used when setting Filters for Library searches or in
the Process Locate dialog.

Flags Flags are a string of one or more characters representing user specific
information. Any characters can be entered, including spaces to a
maximum of eight characters. The order and case (upper or lower) of the
characters are significant.

These values can be used when setting Filters for Library searches, in the
Process Locate dialog or when using the Flagged Entries option in the
Edit Library dialog.

To Enter Textual Data for a User Library Entry

1. Select the Menu Bar Edit, Library command; the Library Edit dialog is invoked.

2. Select the entry whose data is to be entered or modified, and then enter the data.

3. Repeat step 2 as necessary. Each time a new entry is selected, a prompt to save the changes
will be displayed.

4. Select Close to leave the Edit dialog and select Yes to save changes.

Page 11-30
 Library

Indexing a User Library

General

Before a new User Library can be used for Library searching, the Menu Bar Process, Index
Library command must be used to create a Presearch file for the User Library. The Presearch file
contains each Library spectrum reduced to its eight most intense mass-weighted peaks. The Menu
Bar Process, Index Library command invokes the Library Reindex dialog.

Figure 11.26 The Library Reindex dialog

Indexing a Library requires a lot of processing which may take a considerable time, depending on
the size of the Library. The Library Reindex dialog will display an estimate of the time required
to index the Library. Each time new entries are added to the Library, it must be re-indexed before
being used for searching.

To Index a User Library

1. Select the Menu Bar Process, Index Library command; the Library Reindex dialog is
invoked.

2. Select the Start button to start the indexing process. A graphical display displays indexing
progress and gives an indication of the remaining time required. When indexing starts the
Start button changes to a Stop button, the indexing can be aborted at any time by selecting
this.

3. When the indexing is completed, select the OK button and then the Close button to exit.

Page 11-31
Library

Deleting Library Entries

To Delete a User Library Entry

1. Select the Menu Bar Edit, Library command; the Library Edit dialog is invoked.

2. Select the entry to be deleted.

3. Select the Delete Entry button and confirm the deletion with Yes.

Selecting the Edit dialog View button invokes the View dialog. This provides the option to View
Deleted Entries. The User will see the text DELETED above the top left of the spectrum in the
dialog, and all input fields will be grayed. Note also that the Restore Entry button has replaced
the Delete Entry button and can be used to restore this entry. At this point the entry has been
“Flagged as deleted” but has not yet been physically removed from the Library file.

Note:
Only the text associated with a Library entry can be edited. To change the spectrum associated
with a Library entry, delete the entry and then create a new entry by appending the correct
spectrum to the Library.

The Library Locator

General

The Library Locator can be used to look through a Library. Filters can be set up and searches
performed to select certain classes of compounds. The Library Locator dialog is invoked by
selecting the Menu Bar Process, Locate command.

Figure 11.27 The Library Locator dialog

The Library Locator display is similar to the Library Editor display. The Library Locator
display contains the following information about a Library entry: Library Name, Entry No.,
Compound Name, CAS Number, Formula, Molecular Weight, Spectrum and Structure. User
Libraries may also contain Value 1, Value 2 and User Flags.

Page 11-32
 Library

The Locator dialog can be used in two different ways. The first is to select a particular entry for
examination. The second is to set filter parameters that control the entries that Locate will display.

To Select a Particular Entry for Display

1. Either:

Use the arrows to page through the library entries one at a time.

Or:

Select the Select button, type a number into the Entry control and select Update.

2. The Locator dialog display changes to reflect the new selection.

3. Select the Close button to exit.

To Locate Entries with Filters

1. Select the Locator dialog Filters button; the Filters dialog is invoked

2. Set the locate criteria; see below for details.

3. Select the OK button. A message box will display which filters are to be used for the Locate
process.

4. Select the OK button to confirm the criteria.

5. Select Fwd>> or <<Rev to find the next entry matching the locate criteria. Both operations
start at the current entry and either search up in entry number (Fwd), or down (Rev).

6. A message box appears indicating the progress of the location. When the next suitable entry
is found, the display will be refreshed. The Locate process can be aborted by selecting the
Cancel button.

7. The Fwd>> or <<Rev locate processes can be repeated as many times as required.

8. Select Exit to leave the Locator dialog.

To Set the Locate Filters

To set match criteria for the Locate process, select the Locator dialog Filters button; the Filters
dialog is invoked.

The locate filters are set up in exactly the same way as the Library search filters, for more
information see the “Library Search Filters” section, on page 11-13.

Page 11-33
Library

Page 11-34
 Molecular Mass Calculator

Chapter 12 Molecular Mass

Calculator

Page 12-1
Molecular Mass Calculator

Contents
Overview ..................................................................................................................................... 12-3
To Calculate the Molecular Mass for a Given Chemical Formula .............................................. 12-3
Defining User Elements ................................................................................................... 12-4

Illustrations
Figure 12.1 The Molecular Mass Calculator dialog ................................................................... 12-3
Figure 12.2 The User-definable elements dialog........................................................................ 12-4

Page 12-2
 Molecular Mass Calculator

Overview
The MassLynx Molecular Mass Calculator will calculate the average or monoisotopic molecular
mass of any chemical formula.

To Calculate the Molecular Mass for a Given

Chemical Formula
1. Select the MassLynx Tools Shortcut Bar Molecular Weight Calculator icon; the Molecular
Mass Calculator dialog is invoked.

Figure 12.1 The Molecular Mass Calculator dialog

2. Enter the chemical formula using standard IUPAC notation. User-defined elements or
isotopes can be specified by selecting the User elements button, see “Defining User
Elements”, on page 12-4.

3. Choose either Monoisotopic or Average Mass. Monoisotopic mass calculates the mass
using the atomic weight of the most abundant isotope of each element. Average mass
calculates the mass using the average atomic weight of each element taking into account the
relative abundance of its isotopes.

4. Select the +ve or -ve Ion Mode.

5. Enter the range of multiply charged ions to display, i.e. From: 1 To: 4.

6. Select the Calculate button. The calculated mass will appear in the Mass box and the
multiply-charged series in the list box. The current formula can be edited and the mass
recalculated by choosing the Calculate button. The Reset button clears the current formula.

7. The Copy button allows formulae to be copied into the edit control that were previously
pasted into the clipboard from within BioLynx, for example.

Page 12-3
Molecular Mass Calculator

Defining User Elements

1. Select the Molecular Mass Calculator dialog User elements button; the User-definable
elements dialog is invoked.

Figure 12.2 The User-definable elements dialog

2. Enter the required parameters and select the Add button to enter the group in the list. The
Update push-button can be used to edit a particular element or group. Delete removes the
highlighted group in the list box.

3. Up to ten elements, isotopes, molecules can be added to the list.

4. Select the OK button to save the list in the masslynx.ini file for future use.

Page 12-4
 DataBridge

Chapter 13 DataBridge

Page 13-1
DataBridge

Contents
Introduction ................................................................................................................................. 13-3
To Convert a File with DataBridge ............................................................................................. 13-3
General............................................................................................................................. 13-3
Starting the DataBridge Program from the Windows Start Menu.................................... 13-3
Running the DataBridge Program from the MassLynx Sample List................................ 13-6
To Convert an ASCII File to MassLynx Format.............................................................. 13-7

Illustrations
Figure 13.1 The DataBridge dialog ............................................................................................ 13-4
Figure 13.2 The DataBridge - Options dialog ............................................................................ 13-4
Figure 13.3 The Source file select dialog ................................................................................... 13-5
Figure 13.4 The File Information dialog .................................................................................... 13-5
Figure 13.5 The Target directory select dialog........................................................................... 13-6
Figure 13.6 The Header Information dialog ............................................................................... 13-7

Page 13-2
 DataBridge

Introduction
DataBridge is the file conversion program for use with MassLynx; it can perform the following
file conversions:

From: To:
MassLynx NetCDF

MassLynx ASCII

MassLynx Stables OS/2

LAB-BASE MassLynx

NetCDF MassLynx

ASCII MassLynx

PDP11 MassLynx

OPUS MassLynx

Stables OS/2 MassLynx

LAB-BASE Library MassLynx Library

JCAMP Library MassLynx Library

MassLynx Library JCAMP Library

DataBridge allows data to be imported into MassLynx from other sources. This can be LAB-
BASE data, ASCII data, PDP11 data, OPUS Data or data which is in the NetCDF format.
NetCDF is the common data format for mass spectral data specified by the American
Instrumentation Association (AIA). NetCDF allows interchange of mass spectral data from
different manufacturer’s instruments. DataBridge will convert any non-library data in NetCDF
format to MassLynx format for analysis with the MassLynx software.

When converting from PDP11 data, the data must be mass-measured on the PDP11 prior to
conversion, unless the data has been acquired on a TRIO-2 or a 12-250 instrument.

To Convert a File with DataBridge

General

The DataBridge program may be run from the Windows Start Menu, MassLynx Folder (see the
“Starting the DataBridge Program from the Windows Start Menu” section, below). Alternatively,
DataBridge may be selected in the MassLynx Sample List Process column so that it runs when
data acquisition is started, see the “Running the DataBridge Program from the MassLynx Sample
List” section, on page 13-6.

Starting the DataBridge Program from the Windows Start Menu

1. Run DataBridge by selecting the Windows Start button, then selecting Programs,
MassLynx, ; the DataBridge dialog is invoked.

Page 13-3
DataBridge

Figure 13.1 The DataBridge dialog

2. To define the type of conversion required, select the Options button; the DataBridge -
Options dialog is invoked.

Figure 13.2 The DataBridge - Options dialog

3. Select the appropriate file type to convert from, in the Source frame.

4. Select the appropriate file type to convert to, in the Target frame.

5. Select the OK button; the DataBridge - Options dialog is closed.

6. To select the source file, select the Select button; the Source file select dialog is invoked.

Page 13-4
 DataBridge

Figure 13.3 The Source file select dialog

7. Select the file, or files, to be converted.

Note:
Information about a file can be obtained by selecting the File info button. This invokes the File
Information dialog that displays information such as time and date of acquisition, instrument,
number of scans, etc. Select the OK button to close the dialog.

Figure 13.4 The File Information dialog

8. Select the Source file select dialog OK button; the dialog is closed.

Page 13-5
DataBridge

9. To select the directory for the target file, select the Directory button; the Target directory
select dialog is invoked.

Figure 13.5 The Target directory select dialog

10. Select the drive and directory where the converted files are to be saved. DataBridge will
remember the last directory used for each target file type.

11. Select the OK button; the Target directory select dialog is closed.

12. By default the converted file(s) will have the same filename(s) as the original file(s) (but with
the appropriate file extension). If a single source file has been selected, a new name for the
converted file can be entered in the Target frame Filename: text box.

13. Select the Convert button to convert the selected file(s). A scrolling display details the
progress of the conversion.

Note:
If an ASCII file is being converted to MassLynx spectrum format, the Header Information dialog
is invoked, refer to the “Running the DataBridge Program from the MassLynx Sample List”
section, below.

14. Select the Close button to exit DataBridge.

Running the DataBridge Program from the MassLynx Sample List

Note:
Before first running the DataBridge Program from the MassLynx Sample List, the options must be
selected as described in the “Starting the DataBridge Program from the Windows Start Menu”
section, on page 13-3.

1. Double-click on the Process column cell for the sample for which DataBridge is to run; a
drop-down list is invoked.

2. Select the Dbridge option.

3. Start the data acquisition; the DataBridge Program will run as part of the acquisition process,
using the current options.

Page 13-6
 DataBridge

To Convert an ASCII File to MassLynx Format

It is possible to create a single MassLynx format spectrum from an ASCII file. This can be used,
for example, to create a spectrum for a user library. The ASCII file can be created using any plain
text editor, e.g. Windows Notepad.

The ASCII file must contain pairs of mass and intensity values in ascending order from low to
high mass. The values can be separated by any separator, e.g. a TAB character or a comma. The
final value in the file must also be followed by a separator.

When the DataBridge dialog Convert button is selected, the Header Information dialog is
invoked. The information entered here will be displayed when the converted file is selected using
the MassLynx Data Browser dialog, see Chapter 3, “The MassLynx Window and Related
Information”.

To display the spectrum as a continuum spectrum select the Continuum option; leave it deselected
to display the spectrum as a centroid spectrum.

Figure 13.6 The Header Information dialog

Page 13-7
DataBridge

Page 13-8
 AutoLynx

Chapter 14 AutoLynx

Page 14-1
AutoLynx

Contents
Introduction ................................................................................................................................. 14-3
Starting AutoLynx....................................................................................................................... 14-3
AutoLynx Settings....................................................................................................................... 14-4
General............................................................................................................................. 14-4
AutoLynx Settings Dialog: Directories Page ................................................................... 14-4
AutoLynx Settings Dialog: Control Page......................................................................... 14-5
AutoLynx Settings Dialog: Operations page.................................................................... 14-6
AutoLynx Settings Dialog: Results Page ......................................................................... 14-6
Interfacing with External Programs............................................................................................. 14-7
Batch Queuing.................................................................................................................. 14-7
Batch Completion ............................................................................................................ 14-7
Monitoring the Queue Status....................................................................................................... 14-7
Aborting the Queue ..................................................................................................................... 14-7
Accessing Results........................................................................................................................ 14-8
Directory Usage........................................................................................................................... 14-8
File Usage.................................................................................................................................... 14-9
File Structures ............................................................................................................................. 14-9

Illustrations
Figure 14.1 The AutoLynx dialog .............................................................................................. 14-3
Figure 14.2 The AutoLynx Settings Dialog: Directories page ................................................... 14-4
Figure 14.3 The AutoLynx Settings Dialog: Control page......................................................... 14-5
Figure 14.4 The AutoLynx Settings Dialog: Operations page ................................................... 14-6
Figure 14.5 The AutoLynx Settings Dialog: Results page ......................................................... 14-6

Page 14-2
 AutoLynx

Introduction
AutoLynx is an application that enables batches to be submitted to the MassLynx queue for
acquisition, processing and report generation. It allows the number of jobs in the queue to be
monitored. AutoLynx must be running on the same PC as MassLynx, but the Queue and other
directories can be anywhere on the network. Applications can be written (e.g. in Visual Basic)
which create batch files and process the results returned after the data has been acquired and
processed by MassLynx. The application creating the batch files will have to:

• Create batch files in the correct format.

• Write the batch files to the Queue directory.

• Monitor the Status file to determine when a batch has been processed.

• Retrieve processed files from the Processed directory.

• Create an abort file when necessary.

Starting AutoLynx
In Windows Explorer, double click on the file in the main MassLynx directory;
the AutoLynx dialog is invoked.

Note:

Normal Windows procedures may be used to create a Shortcut to the file.

Figure 14.1 The AutoLynx dialog

This dialog displays the batches in the AutoLynx queue, and the date and time they were
submitted. If MassLynx is not running, a message informing the User that MassLynx must be
running to submit batches is also displayed.

Page 14-3
AutoLynx

AutoLynx Settings
General

Select the AutoLynx dialog Settings button to invoke the AutoLynx Settings dialog; this has four
pages:

• Directories.

• Control.

• Operations.

• Results.

AutoLynx Settings Dialog: Directories Page

Figure 14.2 The AutoLynx Settings Dialog: Directories page

Queue Enter the location of the AutoLynx Queue directory.

Processed Enter the location that the batch will be moved to once processing has
been successfully completed. Any results files will also be written to this
directory.

Note:
AutoLynx will create this directory if it does not already exist.

Failed Enter the location that the batch will be moved to if an error has occurred,
or if the batch was aborted.

Note:
AutoLynx will create this directory if it does not already exist.

Page 14-4
 AutoLynx

AutoLynx Settings Dialog: Control Page

Figure 14.3 The AutoLynx Settings Dialog: Control page

Status file Enter the name of the file indicating the current AutoLynx queue status. If
AutoLynx has batches in its queue, this file will exist; it will be deleted
once the queue is empty. By monitoring this file, an external application
can determine when all the batches submitted to the AutoLynx system
have been processed.

Abort file Enter the name of the file that AutoLynx will monitor to determine if the
current batches in the queue should be aborted. If the abort file exists the
current batch will be stopped and all batches in the queue will be moved to
the Failed directory. Any batch file written to the Queue directory when
abort is set will be immediately moved to the Failed directory. Once all
the batches in the queue have been removed the Status file will be deleted.

The external application must create this file to cause an abort, the file
must then be deleted to clear the abort.

Queue Filter Enter the batch file extension type. All files with this extension in the
Queue directory will be added to the AutoLynx queue.

Files of the following formats are supported:

• OpenLynx Batch file (*.olb)

• Comma Separated Value (*.csv).

• Text format (*.txt).

Txt and csv files allow AutoLynx to automatically run the spreadsheet that
would be otherwise submitted to MassLynx by the Menu Bar File, Import
Worksheet command. Refer to the “MassLynx Interfacing Guide” for
further details.

Queue Delay Enter the minimum time, in seconds, between a batch being written to the
Queue directory and it being submitted to MassLynx for processing. This
is intended to ensure that creation of the batch file by the external program
has been completed before the batch is processed.

Page 14-5
AutoLynx

AutoLynx Settings Dialog: Operations page

Figure 14.4 The AutoLynx Settings Dialog: Operations page

Select the options for the types of operation required.

AutoLynx Settings Dialog: Results Page

Figure 14.5 The AutoLynx Settings Dialog: Results page

Print Report Prints the batch results report file. If *.olb files are to be created and
OpenLynx has been specified as the processing type, OpenLynx Browser
*.rpt files are created. The format of this file is defined in the OpenLynx
Browser Report Scheme Settings, refer to the “OpenLynx User’s Guide”
for details.

Create Summary Select this option to create a tab-delimited text file containing a summary
File of the Batch processing results. If OpenLynx is specified as the processing
type, the fields output in the Results Summary file are defined in the
OpenLynx Browser Report Scheme Settings, refer to the “OpenLynx
User’s Guide” for details.

Page 14-6
 AutoLynx

Default Report Enter the name of the OpenLynx Browser report scheme to be used if no
Scheme scheme is defined in the Batch file. If this field is empty, and no scheme is
defined in the batch file, the last scheme selected in the
OpenLynx Browser will be used, refer to the “OpenLynx User’s Guide”
for details.

OpenLynx Enter the location of the OpenLynx Browser program; this will normally
Browser
be in the MassLynx installation directory. Selecting the button will
Location invoke a browser to help locate the required executable file.

Interfacing with External Programs

Batch Queuing

To add batches to the AutoLynx queue, place the relevant batch file in the Queue directory.
AutoLynx displays a list of the batches currently in the queue. Batches will be submitted to
MassLynx for acquisition/processing in the order that they were placed in the Queue directory.

If MassLynx is not running, batches can still be queued, but they will not be processed until
MassLynx is active.

Batch Completion

When a batch has been completed, the batch file, and all other files with the same base name as the
batch, will be moved from the Queue directory. If the batch was completed successfully they will
be moved to the Processed directory; if the batch failed, or an abort was set, they will be moved to
the Failed directory.

Monitoring the Queue Status

The state of the queue can be determined by monitoring the AutoLynx Status file. This file will
only exist if the AutoLynx queue is not empty, or if MassLynx is currently processing a batch; this
file will be deleted once the queue becomes empty and MassLynx is idle. By monitoring this file,
an external process can determine when all the batches submitted to the queue have been run.

Aborting the Queue

An external program can abort all batches in the queue, and stop the acquisition of the current
batch, by creating the Abort file. AutoLynx looks for the Abort file and, if found, all batches will
be removed from the queue. While the Abort file exists, any batch placed in the Queue directory
will be immediately aborted. The Status file will be deleted once all batches have been removed
from the queue and MassLynx is idle. The external program must monitor the Status file and
when this has been deleted, delete the Abort file.

Note:
AutoLynx does not try to open and read the contents of the Abort file.

Page 14-7
AutoLynx

Accessing Results
If the batch was successfully processed and the AutoLynx Settings dialog Results Page Print
Report option was selected, a report will be printed on successful completion of a batch.

Note:
OpenLynx processing must have been performed on the samples in the batch to produce the
necessary OpenLynx Report.

If the batch was successfully processed and the AutoLynx Settings dialog Results Page Create
Summary File option was selected, a file batch_name.txt will have been created in the Processed
directory. This is a tab-delimited text file, the contents of which are dictated by the OpenLynx
Report Scheme Settings used.

Note:
The Results Summary file is produced by the OpenLynx Browser program which requires an
OpenLynx Report file as input (batch_name.rpt). Consequently, OpenLynx processing must have
been performed on the samples in the batch to produce the necessary OpenLynx Report.

Directory Usage
Directory Location Description
Queue User definable in the Settings dialog. Contains all the Batch files in the
current AutoLynx queue. This
Default: C:\AutoLynx directory must exist.

Processed User definable in the Settings dialog. Contains all successfully completed
Batch files and any associated results
Default: C:\AutoLynx\ Processed files. This directory will be created if
it does not exist.

Failed User definable in the Settings dialog Contains all unsuccessful, or aborted,
Batch files and any associated results
Default: C:\AutoLynx\Failed files. This directory will be created if
it does not exist.

Page 14-8
 AutoLynx

File Usage
File Name Description

Status File User definable in the Settings dialog. Exists if AutoLynx is busy, deleted
when all batches have been processed.
Default: C:\Status.sem

Abort File User definable in the Settings dialog. Created by external applications to
abort the AutoLynx queue.
Default: C:\Abort.sem

OpenLynx Batch_name.OLB Describes the samples and processing

Batch file information for a batch. Moved to the
Placed in Queue directory to submit Processed, or Failed, directories after
batch. being processed.

OpenLynx Batch_name.RPT Generated by OpenLynx processing of

Report file the sample. Placed in the Processed
directory upon successful completion.

Results Batch_name.TXT Tab-delimited results file generated by

Summary OpenLynx Browser. The format of
file the file is User-definable through the
Browser Report Schemes Settings.
Placed in the Processed directory upon
successful completion.

File Structures
OpenLynx Batch file: Please contact the Micromass Software Support Department for a copy
of the OpenLynx Batch file structure.

OpenLynx Report file: The fields used, and the order in which they will appear, are defined
using the OpenLynx Browser Report Scheme Settings editor.

Results Summary file: ASCII Tab-delimited file.

The Summary Report format has one line, containing a number of fields, for each sample in the
batch. A single TAB character separates each of the fields.

The fields used, and the order in which they will appear, are defined using the OpenLynx Browser
Report Scheme Settings editor, refer to the “OpenLynx User’s Guide” for further details.

Page 14-9
AutoLynx

Page 14-10
 Accurate Mass Measure

Chapter 15 Accurate Mass Measure

Page 15-1
Accurate Mass Measure

Contents
Introduction ................................................................................................................................. 15-3
Manipulating Files....................................................................................................................... 15-4
To Highlight Files ............................................................................................................ 15-4
To Select Files.................................................................................................................. 15-4
To Deselect Files.............................................................................................................. 15-4
To Change the Output Filename ...................................................................................... 15-4
To Change the Mass Measure Parameters ....................................................................... 15-5
To Change the Mass Filter Parameters............................................................................. 15-5
To Perform Secondary Reference Correction .................................................................. 15-6
To Change the Current Directory..................................................................................... 15-7
Process Type .................................................................................................................... 15-7
Processing Files........................................................................................................................... 15-7
Closing Accurate Mass Measure ................................................................................................. 15-8

Illustrations
Figure 15.1 The Accurate Mass Measure dialog ........................................................................ 15-3
Figure 15.2 The AFAMM - Edit Output File Name dialog........................................................ 15-4
Figure 15.3 The Mass Array Removal From Data Sets dialog................................................... 15-5
Figure 15.4 The Secondary Reference Correction Parameters dialog ........................................ 15-6
Figure 15.5 Invalid data error message ...................................................................................... 15-7

Page 15-2
 Accurate Mass Measure

Introduction
The Accurate Mass Measure (AMM) utility provides a variety of post-acquisition Mass Measure
data processing facilities that can be applied to whole files. AMM is invoked by selecting the
MassLynx Tools Shortcut Bar, Accurate Mass Measure icon.

Figure 15.1 The Accurate Mass Measure dialog

Mass Measure can be performed from within Spectrum in MassLynx, however it can only be done
on a per scan basis. Accurate Mass Measure allows Mass Measure calculations to be applied to all
the scans in all the functions in multiple files in the same directory.

There are also facilities to apply a Secondary Reference Correction or a Mass Filter to files, either
separately or at the same time as applying mass measure. For Secondary Reference Correction
this involves matching the peaks in a data file to the masses in a reference file, correcting the
masses in the data file to reflect those in the reference file, and then writing the corrected masses
to an output file. For Mass Filter, peaks in a data file which match peaks in the reference file are
displayed in a different color when viewed in the Spectrum window.

Page 15-3
Accurate Mass Measure

Manipulating Files
To Highlight Files

A file or files in the Accurate Mass Measure dialog Input File(s) column may be highlighted
using the mouse and conventional Windows techniques.

To Select Files

Select the Accurate Mass Measure dialog Menu Bar Operations, Select command. The will
change to for all the highlighted files.

A single file can be selected by double clicking on the file, or by right-clicking on the file and
selecting the Select command from the invoked pop-up menu.

To select all the files, select the Menu Bar Operations, Select All command.

To Deselect Files

Select the Accurate Mass Measure dialog Menu Bar Operations, Deselect command. The will
change to for all the highlighted files.

A single file can be deselected by double clicking on the file, or by right-clicking on the file and
selecting the Deselect command from the invoked menu.

To deselect all the files, select the Menu Bar Operations, Deselect All command.

To Change the Output Filename

By default, the output filename is input filenameAFAMM.raw.

To change the output name for all files, enter a new name in the Output File Suffix text box and
select the Update button.

To change an individual filename, select the Menu Bar Parameters, Output File command, or
right-click on the filename and select Edit Output File Name from the invoked menu; in either
case, the AFAMM - Edit Output File Name dialog is invoked.

Figure 15.2 The AFAMM - Edit Output File Name dialog

Enter a new name in the text box and select the OK button.

Page 15-4
 Accurate Mass Measure

To Change the Mass Measure Parameters

1. Select Mass Measure or Mass Measure with Peak Filter from the Accurate Mass Measure
dialog Process Type list box.

2. Select at least one file, see the “To Select Files” section, on page 15-4.

3. Select the Menu Bar Parameters, Mass Measure Parameters, Positive Ions or Negative
Ions command; the appropriate Mass Measure dialog is invoked. See the “The Mass
Measure Process” section in Chapter 7, “Spectrum” for details of the parameters.

To Change the Mass Filter Parameters

1. Select Mass Measure with Peak Filter or Peak Filter from the Accurate Mass Measure
dialog Process Type list box.

2. Select the Menu Bar Parameters, Mass Measure Parameters, Mass Filter Parameters
command; the Mass Array Removal From Data Sets dialog is invoked.

Figure 15.3 The Mass Array Removal From Data Sets dialog

3. Select the Browse button and select the required reference file from the invoked dialog.

4. Select Peak Window (ppm) or Peak Window (Da), enter the required window size and
select OK.

The window is ± the entered value (in parts per million or Daltons) about the mass defined in
the reference file.

Page 15-5
Accurate Mass Measure

To Perform Secondary Reference Correction

Mass Measure (of TOF data) allows a lock mass peak to be defined, the Secondary Reference
Correction option allows a file containing more than one peak to be used as reference peaks.

1. Select the Secondary Reference Correction from the Accurate Mass Measure dialog
Process Type list box.

2. Select the Menu Bar Parameters, Secondary Reference Correction Parameters command;
the Secondary Reference Correction Parameters dialog is invoked.

Figure 15.4 The Secondary Reference Correction Parameters dialog

3. Select the Ref File button and select the required reference file from the displayed browser.
Micromass supplied reference files can be found in the c:\masslynx\ref directory.

4. Enter the required parameters, see below.

5. Select the OK button.

Peak Window Enter the range to search the data file for a peak that matches one in the
(ppm) reference file. The window is ± the entered value (in parts per million)
about the mass defined in the reference file, therefore a value of 250 ppm
will result in a search window of 500 ppm.

Intensity Enter the percentage of the most intense peak in the spectrum that a peak
Threshold must be above to be considered as significant. For example, if 10 is
entered, any peak with an intensity of 10% (or more) of the most intense
peak will be considered.

Lock Mass Peaks Enter the percentage of peaks (within the required mass range) in the
Found reference file that must be successfully located in the scan for that scan to
be adjusted for accurate mass.

Largest Peak in This determines how the peak window will be searched. Largest peak in
Window / window uses the mass of the largest peak in the search window. Closest
Closest Peak in peak in window uses the mass of the peak in the data scan closest to that
Window in the reference file.

Page 15-6
 Accurate Mass Measure

To Change the Current Directory

Select the MassLynx Menu Bar File, Open Project, command and select a project from the
displayed browser.

Process Type

The following options are available in the Accurate Mass Measure dialog Process Type list box:

Mass Measure Performs standard Mass Measure processing. For Tof data, lock mass and
dead time correction can be applied by checking the Use TOF mass
correction box on the Mass Measure parameters dialog, selecting the
TOF button and entering the relevant values.

Mass Measure Performs Mass Measurement as above. Additionally, peaks in the

with Peak Filter reference file that match those in the data file are flagged (displayed in a
different color). The flagged peaks will not contribute to the BPI or TIC.

Secondary Applies Secondary Reference Correction to centroid data files. If

Reference continuum data files are selected the data will be Mass Measured first
Correction using the Mass Measure parameters defined within the Accurate Mass
Measure program.

Peak Filter Peaks in the reference file that match those in the data file are flagged
(displayed in a different color). The flagged peaks will not contribute to
the BPI or TIC.

Processing Files
When all the required files have been selected, select the Accurate Mass Measure dialog Process
button. The Process button changes to Cancel, select it to stop processing. The field next to the
Process/Cancel button displays a progress bar to give an indication of the processing time
required.

If the processing parameters are changed and the same file is processed again, a message is
displayed informing the User that the current file name exists; the User is prompted to create a
new file. The name of the new file will be the existing name will a letter appended to it. For
example, processing the Betalac.raw file once will, by default, give an output file named
betalacafamm; if this file is processed a second time, the output file will be named
betalacafamma and, if processed a third time, betalacafammb, etc.

Diode array data and centroided data cannot be processed using this program. If the file contains
continuum data and diode array data then the continuum functions are processed and written to the
output file whereas the diode array functions are just copied. If the file contains only diode array
data, centroided data or diode array and centroided data then the following message is displayed.

Figure 15.5 Invalid data error message

The Status column displays a message if data is valid but cannot be processed, e.g. Lock mass
out of range.

Page 15-7
Accurate Mass Measure

Closing Accurate Mass Measure

To exit the Accurate Mass Measure dialog, select the Exit button, or click on the Windows close
box, at the top right corner of the dialog.

Page 15-8
 IQ Checker

Chapter 16 Installation Qualification

Checker

Page 16-1
IQ Checker

Contents
Introduction ................................................................................................................................. 16-3
Installation Checking................................................................................................................... 16-3
Accessing the IQ Checker; the Installation Qualification Checker Window............................... 16-4
The Installation Qualification Checker Menu Bar....................................................................... 16-4
The File Menu.................................................................................................................. 16-4
The View Menu ............................................................................................................... 16-5
The IQChecker Menu....................................................................................................... 16-5
The Help Menu ................................................................................................................ 16-5
The Installation Qualification Checker Tool Bar ........................................................................ 16-6
Getting Started............................................................................................................................. 16-6
To Open an Existing IQ Check File ................................................................................. 16-6
To Save an IQ Check file ................................................................................................. 16-6
To Change the Font of an IQ Check Report..................................................................... 16-6
IQ Checking ................................................................................................................................ 16-7

Illustrations
Figure 16.1 The Installation Qualification File Checker display................................................ 16-3
Figure 16.2 Installation Qualification Failure Message.............................................................. 16-3
Figure 16.3 The Installation Qualification Checker Window..................................................... 16-4
Figure 16.4 The File Menu......................................................................................................... 16-4
Figure 16.5 The View Menu ...................................................................................................... 16-5
Figure 16.6 The IQChecker Menu.............................................................................................. 16-5
Figure 16.7 The Help Menu ....................................................................................................... 16-5
Figure 16.8 The Select installation directory dialog................................................................... 16-7
Figure 16.9 The Installation Qualification FileChecker dialog .................................................. 16-7
Figure 16.10 Installation Qualification Checker dialog showing typical results ........................ 16-8
Figure 16.11 Installation Qualification Checker dialog showing typical PASS
and FAIL results ................................................................................................... 16-8
Figure 16.12 Installation Qualification Checker dialog showing an internal error message ...... 16-9

Page 16-2
 IQ Checker

Introduction
The Installation Qualification Checker (IQ Checker) program is used to check the validity of a
MassLynx installation.

Immediately after installation, a check is performed automatically to ensure that the files installed
on the User’s PC are the same as those originally packed upon the Installation CD. If any
discrepancies are encountered, an error is displayed and the software should be re-installed.

On successful installation, a file is produced that contains a list of files that have been installed for
the selected configuration. After installation, the current MassLynx installation can be checked
against this file.

Note:
When installing a new version of MassLynx it is recommended that any previous versions are
uninstalled, see the “Uninstalling MassLynx” section, in Chapter 2, “Installing MassLynx”.

After installation of an update to MassLynx, a check is performed, as for the initial installation,
and the file containing the list of installed files is updated to include the new files that have been
installed. Similarly, a check is performed if any new components (e.g. application managers) are
added, or MassLynx is repaired, see the “Repairing MassLynx” section, in Chapter 2, “Installing
MassLynx”.

Installation Checking
The Installation Qualification Checker application is automatically called immediately after
installation. The Installation Qualification FileChecker display is displayed.

Figure 16.1 The Installation Qualification File Checker display

If this check fails, an error message is displayed and the installation is stopped.

Figure 16.2 Installation Qualification Failure Message

Select No and restart the installation.

Page 16-3
IQ Checker

Accessing the IQ Checker; the Installation

Qualification Checker Window
The IQ Checker can be run at any time to verify that the current installation is valid. To access the
IQ Checker after installation, select the Windows Start, Programs, MassLynx, IQ Checker
option; the Installation Qualification Checker Window is invoked.

Figure 16.3 The Installation Qualification Checker Window

The Installation Qualification Checker Menu Bar

The File Menu

Figure 16.4 The File Menu

New Creates a new IQ check file.

Open Opens an existing IQ check file.

Save Saves the current IQ check file to disk.

Page 16-4
 IQ Checker

Save As Saves a copy of the current IQ check file to disk with a new file name.

Print Prints the contents of the current window.

Print Preview Opens a second window to preview the print out.

Print Setup Selects the printer to be used via the standard Windows Printer Setup
dialog.

Recent File Displays the names of up to four most recent IQ check files.

Exit Closes the IQ Checker application.

The View Menu

Figure 16.5 The View Menu

Toolbar Toggles the Tool Bar on and off; a tick signifies “on”.

Status Bar Toggles the Status Bar on and off; a tick signifies “on”.

Fonts Invokes the standard Windows Fonts dialog, allowing the User to select
the font of the output text.

The IQChecker Menu

Figure 16.6 The IQChecker Menu

Run Starts the IQ Checker application, see the “IQ Checking” section, on
page 16-7.

The Help Menu

Figure 16.7 The Help Menu

About Displays the Installation Qualification Checker box, which provides

IQChecker information about the IQ Checker, including the version number.

Page 16-5
IQ Checker

The Installation Qualification Checker Tool Bar

Tool Bar Menu equivalent Purpose
button
File, New Creates a new IQ check file.

File, Open Opens an existing IQ check file.

File, Save Saves the current IQ check file to disk.

File, Print Prints the current window in portrait format.

IQChecker, Run Runs the IQ Checker.

View, Fonts Invokes the standard Windows Fonts dialog,

allowing the User to select the font of the output
text.
Help, About Displays the About Installation Qualification
IQChecker Checker information box.

Getting Started
To Open an Existing IQ Check File

1. Select the Tool Bar button, or select the Menu Bar File, Open command; the Open
dialog is invoked.

2. Select the required IQ Check file (*.iqc).

3. Select the Open button.

To Save an IQ Check file

1. Select the Tool Bar button, or select the Menu Bar File, Save or Save As command; the
Save As dialog is invoked.

2. Enter a name for the new IQ Check file.

3. Select the OK. button.

To Change the Font of an IQ Check Report

1. Select the Tool Bar button, or select the Menu Bar View, Fonts command; the Fonts
dialog is invoked.

2. Select the required Font:, Font style: and Font size:.

3. Select the OK button.

Page 16-6
 IQ Checker

IQ Checking

To start the IQ Checker, select the Tool Bar button, or select the Menu Bar IQ Checker, Run
command. The Installation Qualification Checker, Select installation directory dialog is
invoked.

Figure 16.8 The Select installation directory dialog

By default, the installation directory is c:\masslynx. If the software has been installed in a
different location, enter the new location, or select the Select installation directory button and
select the directory from the displayed browser.

Select the OK button; the IQ Checker checks that the selected directory contains the IQ Check file
containing the list of installed files. If the file is not found an error message is displayed and the
check is aborted.

If the IQ Check file is found, the IQ Checker will check that the file name, create date, file size
and checksum of each *.exe, *.dll, *.hlp and *.cnt file in the selected directory matches those in
the file. The Installation Qualification FileChecker dialog is displayed to indicate the progress
of the check.

Figure 16.9 The Installation Qualification FileChecker dialog

When the check is complete the main window will be updated to display the results.

The first three lines of the report created contain the title, the name of the directory checked, and
the date and time at which the check took place.

After this is a list of files that the IQ checker expects to find in the selected directory, along with
their status (either PASS or FAIL) and nature of failure if appropriate.

A file can fail on one or more of the following criteria:

• File name.

• Create date.

• File size.

• Checksum.

Page 16-7
IQ Checker

Figure 16.10 Installation Qualification Checker dialog showing typical results

Files that pass are displayed in green, those that fail in red. If a file fails on create date, file size
and checksum, the error “Suspected outdated file version” is displayed in blue.

If any of these errors appear the MassLynx directory should be deleted and the software
re-installed.

Figure 16.11 Installation Qualification Checker dialog showing typical PASS and FAIL
results

Page 16-8
 IQ Checker

If an internal error has occurred, the following message will be displayed at the end of the list:

Figure 16.12 Installation Qualification Checker dialog showing an internal error message

This indicates an error in the internal workings of IQ Checker. This error does not necessarily
signify an error with a particular MassLynx setup, but it does mean that the setup cannot be
guaranteed to be correct.

Page 16-9
IQ Checker

Page 16-10
 MassLynx NT Users Guide

Chromatogram
Display Menu, 6-9
Index Edit Menu, 6-8
File Menu, 6-8
Help Menu, 6-14
Menu Bar, 6-8
Process Menu, 6-12
Tool Bar, 6-15
A Tools Menu, 6-14
Window Menu, 6-13
Accurate Mass Measure, 15-1
Chromatogram Display Range dialog,
Accurate Mass Measure dialog, 15-3
6-28
Action On Error options, 4-31
Chromatogram Display View dialog,
Add Library dialog, 11-9
6-32
Adduct Mass, 7-50
Chromatogram Horizontal Axis
AFAMM - Edit Output File Name
Altering the Range, 6-27
dialog, 15-4
Chromatogram Integration, 10-7
Align Chromatogram Time dialog,
Chromatogram Intensity Axis
6-26
Altering the Range, 6-29
Analog Chromatogram dialog, 6-25
Chromatogram Magnify dialog, 6-29
Analysis
Chromatogram Peak Annotation
Starting, 10-20
dialog, 6-35
Append File Select dialog, 11-27
Chromatogram Pointer, 6-27
Append Spectrum dialog, 7-99, 11-27
Chromatogram Tool Bar
Apply Calibration dialog, 7-96
Customizing, 6-16
Auto Find Components dialog, 6-63
Chromatogram Window, 6-7
Auto repair message, 2-11
Chromatograms
AutoLynx, 14-1
Accurate, 6-23
AutoLynx dialog, 14-3
Aligning, 6-26
AutoLynx Settings Dialog
Analog, 6-25
Control page, 14-5
ApexTrack Peak Detection
Directories page, 14-4
Parameters, 6-52
Operations page, 14-6
Automatic Component Finding,
Results page, 14-6
6-64
Automatic Find Components dialog,
Background Subtract, 6-39
7-52
BPI, 6-24
Autosampler Bed Layout dialog, 4-33
Centering the Display, 6-28
Autosampler Bed Layout dialog
Combine Spectra, 6-61
pop-up menu, 4-34
Component Identification, 6-62
Copying To and From the Windows
B Clipboard, 6-75
Deleting Magnification Ranges,
Background Subtract dialog, 6-39,
6-30
7-41
Display Parameters, 6-32
Background Subtract Status dialog,
Displaying, 3-38, 6-19
6-40
Editing Integrated Peaks, 6-54
Backup Scheduler dialog, 2-6
ElectroSpray Data Processing, 6-62
Generating from Spectrum, 6-23
C Integrating, 6-43
Calibration, 7-94 Magnified Ranges, 6-29
Calibration Curve Edit dialog, 10-34 Peak Annotation, 6-34
Calibration Curve Edit, Quantify Peak Lists, 6-69
Compounds? dialog, 10-34 Peak Purity, 6-57
Calibration Curves File, 10-42 Peak Purity Index, 6-59
Calibration Parameters dialog, 7-96 Processing, 6-38
Calibration Report window, 7-95 Processing Multiple, 6-38
Center on Peak List dialog, 6-28 Real-time Display, 6-36
Centre Display dialog, 6-28 Removing from Display, 6-36
Changing the Sample List Format, Restoring the Display, 6-31
4-21

Page I
MassLynx NT Users Guide

Setting the Display Range Defaults, Analog Chromatogram, 6-25

6-31 Append File Select, 11-27
Signal to Noise Ratio, 6-59 Append Spectrum, 7-99, 11-27
Smoothing, 6-41 Apply Calibration, 7-96
Standard Peak Detection Auto Find Components, 6-63
Parameters, 6-47 AutoLynx, 14-3
Summed, 6-23 AutoLynx Settings, 14-4
Text Labels, 6-37 Automatic Find Components, 7-52
TIC, 6-24 Autosampler Bed Layout, 4-33
Closing MassLynx, 3-5 Background Subtract, 6-39, 7-41
Cluster Analysis Options dialog, 8-18 Background Subtract Status, 6-40
Cluster Centroid Options, 8-19 Backup Scheduler, 2-6
Cluster Datafile Options, 8-18 Calibration Curve Edit, 10-34
CODA dialog, 8-21 Calibration Curve Edit, Quantify
CODA Options, 8-21 Compounds?, 10-34
Color dialog, 3-20 Calibration Parameters, 7-96
Colors and fonts Center on Peak List, 6-28
Changing, 3-18 Centre Display, 6-28
Colors and Fonts dialog, 3-18 Chromatogram Display Range,
Combine All Files, 8-26 6-28
Combine All Files dialog, 8-27 Chromatogram Display View, 6-32
File Menu, 8-29 Chromatogram Magnify, 6-29
Operations Menu, 8-29 Chromatogram Peak Annotation,
Combine Datafile Functions dialog 6-35
File Menu, 8-23 Cluster Analysis Options, 8-18
Combine Datafile Functions dialog, CODA, 8-21
8-23 Color, 3-20
Combine Functions, 8-21 Colors and Fonts, 3-18
Combine Spectrum dialog, 7-39 Combine All Files, 8-27
Component Edit, 7-54 Combine Datafile Functions, 8-23
Component Worklist dialog, 6-66 Combine Spectrum, 7-39
Copying a Spreadsheet, 4-13 Component Worklist, 6-66
Create Datafile Map dialog, 9-4, 9-6 Create Datafile Map, 9-4, 9-6
Create Project dialog, 3-34 Create Project, 3-34
Customize Field Display dialog, 4-22 Customize Field Display, 4-22
Customize Toolbar dialog, 6-17, 7-19 Customize Toolbar, 6-17, 7-19
Cut or Delete dialog, 4-28 Cut or Delete, 4-28
Data Browser, 3-27
D DataBridge, 13-4
DataBridge - Options, 13-4
Data Default Chromatogram Range, 6-31
Quantifying, 10-22 Default Spectrum Range, 7-26
Data Acquisition Define Custom Color, 3-21
Starting, 5-3 Diode Array Align, 9-11
Data Browser dialog, 3-27 Display Data, 11-22
Data Dependant Acquisition, 6-74 Display Library Spectrum, 7-99,
Data File Structure, 3-37 11-28
DataBridge, 13-1 Display Quan DB Spectrum, 7-22
DataBridge - Options dialog, 13-4 Display Range, 7-23
DataBridge dialog, 13-4 Display Raw Spectrum, 7-7
Default Chromatogram Range dialog, Edit Component, 7-56
6-31 Edit Components, 7-55
Default Spectrum Range dialog, 7-26 Edit Integrated Peaks, 6-54
Define Custom Color dialog, 3-21 Edit Peak List, 6-70, 11-16
Dialogs Edit Quantify Peak, 10-33
Accurate Mass Measure, 15-3 Edit Text String, 6-37
Add Library, 11-9 EleComp Parameters, 7-78
AFAMM - Edit Output File Name, Enhance Datafile Options, 8-17
15-4 Export SEQUEST compatible file,
Align Chromatogram Time, 6-26 7-35

Page II
 MassLynx NT Users Guide

Fast Centroid, 8-20 MaxEnt 3 (Quad data), 7-73

Field Properties, 4-23 MaxEnt 3 (TOF data), 7-72
File Information, 13-5 MaxEnt 3 Advanced Parameters,
File Open, 6-71 7-73
Filters, 11-13 MaxEnt Reconstruction status, 7-70
Font, 3-19 MaxEnt3 Sequence status, 7-74
Format, 3-41 Modify Calibration, 7-97
Format DB List, 10-30 Molecular Mass Calculator, 12-3
Format DB List, 11-19 New Chromatogram, 6-20
General Method, 10-15 New Spectrum, 7-22
Get Peak List Entry, 6-73 Number Of Injections, 4-30
Header Editor, 3-39 Open File, 4-12
Header Information, 13-7 Options, 3-23
History Selector, 3-29, 7-37 Options, Multi-probe, 3-25
Import Data, 4-18 Output File, 8-16, 8-26
Import Worksheet, 4-14 Paste Area or Samples, 4-29
Input Datafile, 8-13, 8-24 Paste Option, 4-29
Installation Qualification Checker, Peak Detect, 6-47, 7-49
16-8 Peak Purity, 6-57
Installation Qualification Periodic Table, 7-89
FileChecker, 16-7 Print, 3-43
Integrate chromatogram, 6-44, Print Report, 10-37
10-17 Process, 5-6
Integration Window, 10-17 Program Maintenance, 2-9
Isotope Cluster Parameters, 7-80 Quantify Chromatogram Display,
Isotope modelling, 7-76 10-39
Isotopes, 7-90 Quantify Display, 10-24
Library Compare, 11-25 Quantify Method Editor, 10-12
Library Data Browser, 11-10 Quantify Process, 10-35
Library Display View, 11-17 Quantify Report Format, 10-38
Library Edit, 11-29 Quantify Reports, 10-36
Library Locator, 11-32 Quantify Samples, 10-22
Library Reindex, 11-31 Queue Properties, 5-7
Library Search List, 11-8 Real Number Field Properties, 4-23
Library Search Parameters, 11-11 Refine Spectrum, 7-38, 11-24
List Field Justification, 10-31, Remove Chromatogram, 6-36
11-20 Remove Spectra, 7-33
Load Sample List Format, 4-20 Report Table Format, 10-26
Lock Mass, 7-97 Response Threshold, 6-53, 10-18
Make new calibration, 7-95 Retention Index Status, 6-14
Make Retention Index Calibration, Retention Index Table, 6-76
6-77 Samples, 4-13, 4-27
Manual Find Components, 7-51 Save As, 4-12
Map Data Browser, 9-5 Save Sample List Format, 4-21
Map Display Ranges, 9-8 Scan Select, 11-11
Map Intensity Scaling, 9-8 Secondary, 10-13
Map View, 9-9 Secondary Reference Correction
Mass, 7-79 Parameters, 15-6
Mass Array Removal From Data Select Entry, 10-29
Sets, 15-5 Select File, 4-24
Mass Chromatogram dialog for SIR Select Hot Symbol, 7-88
data, 6-21 Select installation directory, 16-7
Mass Chromatogram for full scan Select Mass, 9-12
data, 6-21 Select Project, 3-35
Mass measure, 7-46 Select Raw Spectrum, 7-7
MassLynx Options, MassLynx Select Time, 9-12
Installation directory, 2-6 Select Wavelength, 9-12
MassLynx Security Options, 2-5 Set Adduct Mass, 7-50
MaxEnt 1, 7-60 Short Cut Remove Chromatogram,
MaxEnt 2, 7-69 6-36

Page III
MassLynx NT Users Guide

Short Cut Remove Spectrum, 7-33 Elemental Composition Window,

Signal to Noise, 6-60 7-80
Smooth chromatogram, 6-42, 10-18 Edit Menu, 7-82
Source file select, 13-5 File Menu, 7-82
Spectrum Center, 7-44 Help Menu, 7-83
Spectrum Data Browser, 7-37 Menu Bar, 7-82
Spectrum Display, 7-28 Process Menu, 7-83
Spectrum Magnify, 7-24 Tool Bar, 7-84
Spectrum Peak Annotation, 7-31 View Menu, 7-83
Spectrum Print Report, 7-27 Enhance Datafile Options, 8-17
Spectrum Real-Time Update, 7-34 Enhance Datafile Options dialog,
Spectrum Save, 7-36 8-17
Spectrum Smooth, 7-42 Experimental Record
Start Sample List Run, 5-3, 10-21 Window Options Menu, 3-32
Strip Data Browser, 8-14, 8-25 Experimental Record Window, 3-31
Strip Datafile, 8-6 File Menu, 3-32
Subtract Background File, 8-15 Export SEQUEST compatible file
Subtract Datafile Options, 8-16 dialog, 7-35
Subtract Hit, 11-26
Summary Information, 4-21 F
Superatom Limits, 7-93
Superatom Tables, 7-92 Fast Centroid dialog, 8-20
Target directory select, 13-6 Field Properties dialog, 4-23
TIC Chromatogram, 6-24 File Conversion with DataBridge,
TOF Accurate Mass, 7-47 13-3
TOF Transform, 7-48 File Information dialog, 13-5
Transform, 7-58 File Open dialog, 6-71
Uninstall Option: Uninstall Java Filters dialog, 11-13
components, 2-11 Font Dialog, 3-19
User Text, 3-40 Format DB List dialog, 10-30, 11-19
User-definable elements, 7-77, 12-4 Format dialog, 3-41
Diode Array Align dialog, 9-11
Diode Array Data G
Displaying, 9-4, 9-10
Directory Structure, 3-35 General Method dialog, 10-15
Display Data dialog, 11-22 Get Peak List Entry dialog, 6-73
Display Library Spectrum dialog,
7-99, 11-28 H
Display Quan DB Spectrum dialog,
7-22 Header Editor, 3-39
Display Range dialog, 7-23 Header Editor dialog, 3-39
Header Information dialog, 13-7
Help, 3-44
E History Selector dialog, 3-29, 7-37
Edit Component dialog, 7-56 Hot Symbol
Edit Components dialog, 7-55 Select, 7-89
Edit Integrated Peaks dialog, 6-54
Edit Peak List dialog, 6-70, 11-16 I
Edit Quantify Peak dialog, 10-33
Edit Text String dialog, 6-37 Import Data dialog, 4-18
EleComp Parameters dialog, 7-78 Import Worksheet dialog, 4-14
ElectroSpray Data Processing, 7-49 Importing a Worksheet, 4-14
Element Importing Data, 4-17
Deselect, 7-90 Input Datafile dialog, 8-13, 8-24
Select, 7-89 Installation Qualification Checker
Elemental Composition, 7-78 dialog, 16-8
Elemental Composition Parameters, Installation Qualification Checker
7-84 Window, 16-4
Elemental Composition warning box, Installation Qualification Failure
7-93 Message, 16-3

Page IV
 MassLynx NT Users Guide

Installation Qualification File Library Spectra

Checker display, 16-3 Manipulating, 7-99
Installation Qualification FileChecker List Field Justification dialog, 10-31,
dialog, 16-7 11-20
Installing MassLynx Libraries, 2-9 List Properties Window, 3-8
Integrate chromatogram dialog, 6-44, Load Sample List Format dialog,
10-17 4-20
Integration Window Dialog, 10-17 Lock Mass dialog, 7-97
IQ Checker, 16-1
File Menu, 16-4 M
Help Menu, 16-5
IQChecker Menu, 16-5 Make new calibration dialog, 7-95
View Menu, 16-5 Make Retention Index Calibration
Isotope dialog, 6-77
Deselect, 7-90 Manual Find Components dialog,
Select, 7-89 7-51
Isotope Cluster Abundance Plots, Manual Peak Integration, 10-32
7-75 Map, 9-1
Isotope Cluster Parameters dialog, Map Data Browser dialog, 9-5
7-80 Map Display Ranges dialog, 9-8
Isotope modelling dialog, 7-76 Map Intensity Scaling, 9-8
Isotopes dialog, 7-90 Map Intensity Scaling Dialog, 9-8
Map Tool Bar, 9-6
Map View dialog, 9-9
L
Mass Array Removal From Data Sets
Library, 11-1 dialog, 15-5
Append File Select prompt, 11-27 Mass Chromatogram dialog for full
Append Spectrum prompt, 11-27 scan data, 6-21
Compare, 11-25 Mass Chromatogram dialog for SIR
Delta Window, 11-22 data, 6-21
Hit List Window, 11-18 Mass Measure, 7-46
Hits Window, 11-21 Mass measure dialog, 7-46
Indexing a User Library, 11-31 MassLynx Bar, 3-11
Mainsearch, 11-7 Displaying, 3-12
Presearch, 11-6 MassLynx Folder, 2-7
Printing Results, 11-23 MassLynx Menu Bar, 3-7
Refining the Search Spectrum, File Menu, 3-7
11-24 Help Menu, 3-10
Search, 11-6 Run Menu, 3-9
Search Filters, 11-13 Security Menu, 3-9
Search List, 11-8 View Menu, 3-9
Search Parameters, 11-11 MassLynx Options, MassLynx
Search Results, 11-16 Installation directory dialog, 2-6
Search Spectrum, 11-10 MassLynx Security Options dialog,
Structure Window, 11-23 2-5
Subtract, 11-26 MassLynx Tool Bar, 3-10
Tool Bar, 11-6 MassLynx Window, 3-6
User Libraries, 11-26 MassLynx Wizard Welcome Screen,
Window, 11-5 2-4
Library Compare dialog, 11-25 MassLynx.ini file, 3-26
Library Data Browser dialog, 11-10 MaxEnt 1, 7-59
Library Display View dialog, 11-17 MaxEnt 1 dialog, 7-60
Library Edit dialog, 11-29 MaxEnt 2, 7-69
Library Locator, 11-32 MaxEnt 2 dialog, 7-69
Library Locator dialog, 11-32 MaxEnt 3, 7-71
Library Reindex dialog, 11-31 MaxEnt 3 Advanced Parameters
Library Search List dialog, 11-8 dialog, 7-73
Library Search Parameters dialog, MaxEnt 3 dialog (Quad data), 7-73
11-11 MaxEnt 3 dialog (TOF data), 7-72

Page V
MassLynx NT Users Guide

MaxEnt Reconstruction status dialog, Peak List File

7-70 Creating, 6-71
MaxEnt3 Sequence status dialog, Opening, 6-72
7-74 Peak Lists File, 10-42
Menus Peak Purity dialog, 6-57
Autosampler Bed Layout dialog Peak Thresholding, 10-18
pop-up, 4-34 Periodic Table dialog, 7-89
Modify Calibration dialog, 7-97 Pointer, 6-27
Molecular Mass Calculator, 12-1 Print dialog, 3-43
Molecular Mass Calculator dialog, Print Report dialog, 10-37
12-3 Printing Data, 3-42
Moving Mean, 10-18 Process dialog, 5-6
MS Status Bar, 3-17 Processed Data Labels, 3-29
Program Maintenance dialog, 2-9
N Projects, 3-33, 10-10
Creating, 3-33
NetCDF, 13-3 Opening, 3-35
New Chromatogram dialog, 6-20
New Features in MassLynx NT 3.0,
Q
1-36
New Features in MassLynx NT 3.1, QTOF Accurate Mass, 7-47
1-35 Quantification
New Features in MassLynx NT 3.2, Export to LIMS File, 10-39
1-33 Quantify, 10-1
New Features in MassLynx NT 3.3, Files, 10-41
1-31 Tool Bar, 10-23
New Features in MassLynx NT 3.4, Quantify Chromatogram Display
1-28 dialog, 10-39
New Features in MassLynx NT 3.5, Quantify Display dialog, 10-24
1-22 Quantify Graphs Window, 10-28
New Features in MassLynx NT 4.0, Quantify Manager Bar, 10-5
1-4 Quantify Method, 10-11
New Spectrum dialog, 7-22 Adding a New Compound, 10-19
Number Of Injections dialog, 4-30 Creating, 10-19
Deleting a Compound, 10-20
O Deleting All Compounds, 10-20
Inserting a New Compound, 10-19
Open File dialog, 4-12 Modifying an Existing Compound,
Opening MassLynx, 3-5 10-20
Options dialog, 3-23 Propagating General Parameters to
Options, Multi-probe dialog, 3-25 All Compounds, 10-19
Output File dialog, 8-16, 8-26 Propagating Integration Parameters
to All Compounds, 10-19
P Selecting an Existing Method,
10-19
Parameters Quantify Method Editor dialog,
Elemental Composition, 7-84 10-12
Parameters dialog Quantify Method File, 10-42
General Parameters page, 7-85 Quantify Method Parameters, 10-12
Symbol Parameters page, 7-87 Quantify Method Peak Integration
Paste Area or Samples dialog, 4-29 Parameters, 10-16
Paste Option dialog, 4-29 Quantify Peak List Window, 10-30
Peak Detect dialog, 6-47, 7-49 Quantify Process dialog, 10-35
Peak Edit Error warning Quantify Report Format dialog,
Peak can’t overlap other peaks, 10-38
6-56 Quantify Reports
Start and end positions are invalid, Printing, 10-36
6-56 Quantify Reports dialog, 10-36
Peak Integration in Progress message, Quantify Results
6-46 Changing, 10-32

Page VI
 MassLynx NT Users Guide

Quantify Samples dialog, 10-22 Selecting Areas of the Display,

Quantify Summary Window, 10-25 4-23
Quantify Window, 10-23 Sorting Rows, 4-30
Quantifying Data, 10-22 Specifying the Number of
Queue Injections, 4-30
Job Properties, 5-6 Updating from the AutoSampler
Properties, 5-7 Bed Layout, 4-32
Queue Bar, 3-16, 5-5 Samples dialog, 4-13, 4-27
Queue Properties dialog, 5-7 Samples, Clear sub-menu, 4-8
Samples, Column sub-menu, 4-8
R Samples, Fill sub-menu, 4-7
Samples, Format sub-menu, 4-9
Real Number Field Properties dialog, Samples, Sort sub-menu, 4-10
4-23 Save As dialog, 4-12
Refine Spectrum dialog, 7-38, 11-24 Save Sample List Format dialog,
Remove Chromatogram dialog, 6-36 4-21
Remove Spectra dialog, 7-33 Savitzky Golay, 10-18
Repairing MassLynx, 2-11 Scan Select dialog, 11-11
Report Table Format dialog, 10-26 Secondary dialog, 10-13
Response Threshold dialog, 6-53, Secondary Reference Correction
10-18 Parameters dialog, 15-6
Retention Index, 6-76 Select Entry dialog, 10-29
Calibration, 6-77 Select File dialog, 4-24
Checking Calibration Status, 6-79 Select Hot Symbol dialog, 7-88
Displaying Values, 6-78 Select installation directory dialog,
Retention Index calibration curve, 16-7
6-78 Select Mass dialog, 9-12
Retention Index Status dialog, 6-14 Select Project dialog, 3-35
Retention Index Table Select Time dialog, 9-12
Deleting, 6-77 Select Wavelength dialog, 9-12
Setting up, 6-77 SEQUEST Files
Retention Index Table dialog, 6-76 Exporting, 7-34
Set Adduct Mass dialog, 7-50
S Short Cut Remove Chromatogram
dialog, 6-36
Sample List, 4-1, 10-10 Short Cut Remove Spectrum dialog,
Sample List File, 10-41 7-33
Sample List Formats, 4-19 Shortcut Bar, 3-12
Sample List Menu Bar, 4-6 Application Managers, 3-16
Edit Menu, 4-6 Instrument, 3-12
Samples Menu, 4-7 Tools, 3-14
Sample Lists Signal to Noise dialog, 6-60
Action On Error, 4-31 Smooth chromatogram dialog, 6-42,
Adding Samples, 4-27 10-18
Changing the Column Width, 4-21 Smoothing, 10-17
Creating, 4-13 Source file select dialog, 13-5
Deleting Data, 4-28 Spectra
Displaying Specific Columns, 4-21 Background Subtract, 7-40
Editing Data, 4-24 Combine, 7-39
Editing Data in an Area, 4-26 Copying, 7-97
Editing Field (Column) Properties, Displaying, 3-37
4-22 Integrating, 7-48
Inserting Rows, 4-26 Refine, 7-38
Loading an Existing Format, 4-20 Smooth, 7-41
Opening, 4-11 Spectrum, 7-1
Printing Files, 4-13 Adding or Replacing Spectra, 7-21
Removing a Column, 4-22 Altering the Range of Both Axes,
Saving, 4-12 7-23
Saving a Format, 4-20 Altering the Range of the Intensity
Axis, 7-23

Page VII
MassLynx NT Users Guide

Altering the Range of the Mass Superatom

Axis, 7-23 Database, 7-92
Center, 7-43 Limits, 7-93
Deleting Magnification Ranges, Tables, 7-91
7-25 Superatom Limits dialog, 7-93
Display Menu, 7-10 Superatom Tables dialog, 7-92
Displaying as a List, 7-26 Survey Spectrum, 7-62
Edit Menu, 7-9 System Global Parameters, 3-22
File Menu, 7-9 System Requirements, 2-3
Help Menu, 7-17
Menu Bar, 7-9 T
Process Menu, 7-13
Processing, 7-35 Target directory select dialog, 13-6
Restoring the Display, 7-25 Text Files
Setting Magnified Ranges, 7-23 Creating Import Data, 4-19
Setting the Display Range Defaults, Text Labels
7-26 Spectrum, 7-34
Tool Bar, 7-17 The Display Raw Spectrum dialog,
Tools Menu, 7-15 7-7
Window, 7-8 The Mass dialog, 7-79
Window Menu, 7-16 The Select Raw Spectrum dialog, 7-7
Spectrum Center dialog, 7-44 TIC Chromatogram dialog, 6-24
Spectrum Data Browser dialog, 7-37 TOF Accurate Mass dialog, 7-47
Spectrum Display dialog, 7-28 TOF Transform dialog, 7-48
Spectrum Display Parameters, 7-28 Tool Bar
Spectrum Magnify dialog, 7-24 Adding Buttons, 6-18, 7-20
Spectrum Peak Annotation dialog, Changing the Display Order, 6-18,
7-31 7-21
Spectrum Print Report dialog, 7-27 Removing, 6-19
Spectrum Real-Time Update dialog, Removing Buttons, 6-18, 7-20
7-34 Resetting, 6-19, 7-21
Spectrum Save dialog, 7-36 Tool Bars
Spectrum Smooth dialog, 7-42 Spectrum, 7-17
Spectrum Tool Bar Transform, 7-58
Customizing, 7-18 Component Find, 7-50
Spectrum Window, 7-8 Transform dialog, 7-58
Start Sample List Run dialog, 5-3,
10-21 U
Starting Data Acquisition, 5-3
Strip, 8-5 Uninstall Option: Uninstall Java
Cluster, 8-5 components dialog, 2-11
CODA, 8-5 Uninstalling MassLynx, 2-10
CODA data file, 8-12 Update Disks, 2-8
Clustered data file, 8-11 User Text dialog, 3-40
Enhanced data file, 8-8 User-definable elements dialog, 7-77,
Subtracted data file, 8-10 12-4
Enhance, 8-5
Subtract, 8-5 W
Strip Data Browser dialog, 8-14, 8-25
Strip Datafile dialog, 8-6 Window Commands, 3-44
File Menu, 8-7
Help Menu, 8-8
Options Menu, 8-7
Subtract Background File dialog,
8-15
Subtract Datafile Options, 8-16
Subtract Datafile Options dialog, 8-
16
Subtract Hit dialog, 11-26
Summary Information dialog, 4-21

Page VIII