Week10 PracticalHoms Compulsory aes PranicalReport20

UTARHospitalVisit Prounical
Assessment 4
Pre labRequired Coursework25

Clinical Biochemistry
Introduction to clinical biochemistry
laboratory
Topic6 BiotatPrinciples
7 11 4 Dr Chew Choy Hoong
Test2 Friday chewch@utar.edu.my
06092024 12.51 ImportantTests
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 Learning objectives

To understand:

✓ how sample handling, analytical and biological factors

can affect test results in health and disease and how these
relate to the concept of a test reference range;

✓ the concepts of accuracy, precision, test sensitivity, test

specificity in the quantitative assessment of test
performance.

normalreferencerange
dolferswithdifferenthospitals
laboratory
 Clinical Biochemistry/
Medical Biochemistry/ Clinical
Chemistry/ Chemical Pathology ?
A branch of the study of laboratory medicine in which
chemical and biochemical methods are applied to the
study of disease

cuesare me norm
Automated
q

Most laboratories are now

computerised, and the use of
bar-coding of specimens
and automated methods of
analysis allows a high degree
of productivity and improves
the quality of service.

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IN
UTAR!!!!!!!
Clinical Laboratory Organisation

Core Laboratory (73% of volume)

Chemistry, haematology, toxicology, immuno-
chemistry, special chemistry, coagulation, urinalysis

Microbiology (16% of volume)

Blood/urine cultures, serology
from
car wifi
Transfusion Services (3% of volume)
Blood typing and cross-matching
 Types of Chemistry

Electrolytes, glucose, blood urea nitrogen (BUN),

creatinine, phosphate, total protein, bilirubin,albumin liver
magnesium, calcium, cholesterol, triglycerides, etc.
kidneys

Blood gases: pO2, pCO2, pH, calc. Parameters

insulin GHlevels
Immunochemistry: endocrine, specific protein, tumour
markers

effects
Toxicology: Therapeutic Drug Monitoring (TDM)

Urinalysis
PATIENT
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mutations
gene

caissuestain
tissues

blood
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emiabase
balance
 Uses of Biochemical Testing
Diagnosis Prognosis severity

Information on the likely of

diagnosed
Confirmation/rejection condition
of clinical diagnosis outcome of disease

Functions

Monitoring Screening EE iiiiwrauine

Monitoring progression or Detection of subclinical preventcomplication
accumulation
before
response to treatment disease
screening
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markers
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Antigenpresencelevel
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monitoring monitor
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 Diagnosis

History and Physical Examination of patient --form

Differential Diagnosis i.e. Hypothesis
arenotperfectaccuracy errorsduringbefore after
Lab tests and Radiology to support or reject hypothesis.
Limitations of tests must be taken into account.

Interpretation must be carefully done in context of clinical

details
 Prognosis
Prognosis
Serial tests to identify progressive disease. For example:
creatinine in renal failure; used to indicate when dialysis
may be required

Tests to identify risks of disease in future in certain groups

cholesterol in “at risk families”

Likely outcome of disease

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somecansare untestablebeforetreatmentEglipemicbrood
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impute
 Monitoring

To monitor progression of illness

- Follow natural history of disease

To monitor effectiveness of treatment

- i.e. glucose levels in diabetes mellitus in response to insulin
treatment.
- Development of complications side effects of treatments (i.e.
hypokalaemia during treatment of diuretics)
EIE.EEIifiii c nypereeni.iii iii in
cardiacfailure
Drug Toxicity / Therapeutic Drug Monitoring (TDM)
 Screening
Screening
Determine if a condition is present subclinically
- i.e. Neonatal screening phenylketonuria (PKU).

Natural history known Topic9

L

Ease of treatment

Positive outcome
 Clinical Biochemistry
Requesting and Interpreting Tests
 Sampling

Test Request e.g test

blood

Clinician requests that analysis be conducted.

Requirements DOB Age

1 ggfrange
Name, DOB, Gender, Hospital No.
Ward, Address
Requesting Clinician
emeofinterventionbymeaicalseat f.it
Hypothesis / Problem
Tests sought, time of sampling, date fi ii ionaf
blood

Clinical details, Drugs or Therapy. pities

Keleters
mustbeorderedbydoctors noted
 DIFFTEST DIFF collection tube A bloodsugarlevel
e.gHaema EDTAtube commonly EYE muggins

PATIENT”S
PARTICULARS

TEST REQUIRED
 Sampling Issues

Plasma, serum (Mostly)

Cerebrospinal fluid, pleural fluid, urine, etc
Bottle and preservative, anti-coagulant used
Age of sample
Time of sampling
Guidance from lab on sample requirements
Correct label
Transport
High risk specimens
 Plasma vs Serum
Plasma is obtained by collecting
blood into an anticoagulant and
separating the fluid, plasma
phase from the blood cells by
centrifugation.

Serum is the corresponding fluid

phase when blood is allowed to
clot.
 Near Patient Testing/ Point of
Care Testing

Instruments are available that can perform certain tests at

remote locations, such as at the bedside on in a clinical
care unit
– Blood glucose
– Urinalysis
– Blood gases
– Electrolytes/BUN/Creatinine
– Cardiac markers (Troponin I & T, CK-MB)

Near-patient tests are nearly always more expensive, than

the same tests performed in the central laboratory
 Reasons for Near-Patient
Testing/Point of Care

Tests are of urgent importance, and results will affect the

immediate management of the patient
– Blood gases, electrolytes

Tests are so common, simple and cheap that it is more

economical to perform them at the point of care (home)
– Blood glucose, urinalysis
Advantages and Disadvantages of Point-
of-Care Testing (POCT)
The CoaguChek® XS Plus
uses new technology for the
determination of INR
(international normalised
ratio) - values (or the level of
prothrombin) of the patients
blood.

Urinalysis
Urinalysis BloodGlucose
glucosemeter
monitoringArterialBlood
Blood gas
Gasanalyser
Analyser
Steps in obtaining a laboratory test
 Sources of error in laboratory
results

 Pre-analytical
Most frequent source of errors (up to 70%).

 Analytical
Infrequent in laboratory tests, however may be higher in POCT due to non-lab trained
personnel operating devices.

 Post-analytical
Second most common among laboratory-based results
Sources of Error
An example of Interfering
Substances
 Pre-analytical errors

Collection
– Was the right tube used?
– Was venipuncture performed correctly?
– Was the specimen properly stored?

Identification
– Was the blood collected from the correct patient?
– Was the blood correctly labeled?
Patient name, ID, date, time of collection, phlebotomist
Blood collection tubes
Blood collection tubes for specific
biochemical tests
 Collection tubes

Red-top tubes contain no anticoagulants or preservatives

Red-top tubes are used for collecting serum

– 10-15 minutes is required to allow blood to clot before
centrifuging
– Used for blood bank specimens, some chemistries
 Collection tubes

Gold (and “tiger”) top

tubes contain a gel
that forms a physical
barrier between the
serum and cells after
centrifugation

No other additives are

present

Gel barrier may affect

some lab tests
 Collection tubes

Gray-top tubes contain either:

– Sodium fluoride (and potassium oxalate), or
– Sodium iodoacetate (and heparin)

Both perservatives stabilize glucose in plasma

– NaF/oxalate inhibits enolase
– Iodoacetate inhibits glyceraldehyde-3-phosphate
dehydrogenase
 Collection tubes

Green-top tubes contain either the sodium or lithium salt of

heparin
– The amount of Na+ or Li+ is insignificant

Heparin inhibits thrombin, so blood does not clot (plasma)

The advantage of plasma is that no time is wasted waiting

for the specimen to clot
 Collection tubes

Lavender-top tubes contain EDTA, which binds calcium

and inhibits coagulation

Used for haematology, and some chemistries (i.e.

lipids/lipoprotein)
 Collection tubes

Blue-top tubes contain sodium citrate, which binds calcium

and inhibits coagulation

The blood/anticoagulant ratio must be precisely known,

since the tubes are used for coagulation studies.
 Collection tubes

Brown and Royal Blue top tubes are specially cleaned for
trace metal studies

– Brown-top tubes are used for Plumbum analysis

– Royal blue-top tubes are used for other trace element
studies (acid washed)
 Specimen identification

One of the commonest sources of error in lab results is

misidentified specimens

The blood bank has stricter requirements for specimen

identification
 How biochemical results
are expressed
Most biochemical analyses are quantitative, although simple
qualitative or semi-quantitative tests, such as those for the
presence of glucose in urine, are commonly encountered
methods used for point of care testing.

Many tests measure the amount of the analyte in a small

volume of blood, plasma, serum, urine or some other fluid or
tissue.

Results are reported as concentrations, usually in terms of the number

of moles in one litre (mol/L). Enzymes are expressed as enzyme activity
in ‘units’. Some hormone measurements are expressed as ‘units’. Large
molecules such as proteins are reported in mass units (grams or
milligrams) per litre. Blood gas results (PCO2 or PO2) are expressed in
kilopascals (kPa),
The Normal Reference Range for Analytes
 Variation in results
Biochemical measurements vary for two reasons:

Analytical variation is a function of analytical

performance.
– precision and accuracy
– sensitivity and specificity
– quality assurance
– reference intervals.

Biological variation is related to the actual changes that

take place in patients’ body fluids over a period of time.
 Precision and Accuracy
Precision is the reproducibility of an
analytical method.
Accuracy defines how close the
measured value is to the actual
value.

Precision is how closely repeated

measurements match each other.

Accuracy is how closely a measurement

matches the correct or expected value.
Which method is more precise or accurate?
 Analytical sensitivity and specificity
Sensitivity is the ability of a method to detect small quantities of
measured analyte.

Specificity of the ability of the method to measure only one

analyte for which it has been designed and as an indicator of
the method’s freedom from interference.
Quality assurance
Laboratory staff monitor performance of assays using quality
control samples to give reassurance that the method is
performing satisfactorily with the patients’ specimens.

Internal quality control samples are analysed regularly.

Which method is more sensitive or specific?

ROC curves (receiver

operating characteristic
curves) for three
hypothetical tests
 even itfalls Reference intervals
elweenmenormalrange
vdoesnotparticularly thatthepatientisfreefrom
means negatived
Biochemical test results are
usually compared to a
reference interval chosen
arbitrarily to include 95% of
the values found in healthy
volunteers.

This means that, by definition,

5% of any population will
have a result outside the
reference interval.
 Specificity and sensitivity of tests

(The use of the terms specificity and sensitivity in this context

should not be confused with the same terms used to describe
analytical performance).

The specificity of a test measures how commonly negative

results occur in people who do not have a disease.

Sensitivity is a measure of the incidence of positive results in

patients who are known to have a condition.

An ideal diagnostic test would be 100% sensitive, showing

positive results in all diseased subjects, and 100% specific,
with negative results in all persons free of the disease.
 diseases
Sensitivity identify
non diseasedpatients
Specificity identify

When a test has a sensitivity of 80%, it can correctly

identify 80% of people who have the disease, but it
misses 20%. This smaller group of people have the
disease, but the test failed to detect them—this is
known as a false negative.

A test that has an 80% specificity can correctly identify

80% of people in a group that do not have a disease,
but it will misidentify 20% of people. That group of 20%
will be identified as having the disease when they do
not, this is known as a false positive.
 O

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 Biological factors affecting the
interpretation of results
Age
44L B P.int n tiue
Gender
Diet - Fasting/Fed glucose triglycerides
Posture – Redistribution of fluid may affect the result iii piins
Physiological State – stress/anxietyLimit's.inorepinepwinesfEpig eana
Pregnancyamatemalnormo magician
Exercise
Lui
Timing – variations in day/night/menstrualegckitsii
cyclehormones
timeofnfea.fi
morning
udiaemankersf
Medical history - Infection and/or tissue injury can affect
biochemical values independently aftertan
Drug history - Drugs may have specific effects on the
plasma concentration of some analytes.
Diuretics Potassium Lamond
 Other factors affecting the
interpretation of results

Plasma/Serum differences totalprotein

 Plasma>Serum (TP, Ca2+)
 Plasma<Serum (CK, K+, PO4-)
 Supine vs. sitting or standing
 Supine patient: decrease in TP, albumin, lipids, Ca
 Haemolysis
 Recent strenuous exercise
 Decreases: lipid, K+
 Increases: CK, AST, ALT, creatinine on

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Analytes significantly affected

Be

by haemolysis
Total protein, albumin, lipids, iron, calcium, bilirubin,
cholesterol, triglycerides, glucose, norepinephrine,
wz y altered
heavily
potassium, magnesium, phosphate, and enzymes
(alkaline phosphatase, aspartate transaminase)

maffarance

Hemolyzed/Haemolysed
ie Lipemic/ Normal
Lipaemic serum
blood serum
triglyceride
nign cholesterol
nottotal
Icteric – Lipaemic - haemolytic –
Jaundiced – excess High lipid (triglyceride) haemolyzed blood,
bilirubin haemoglobin released from
damaged RBCs

haemolyzedbrood
strawberry
milk lipenicblood
 Case study
premium

A blood specimen was taken from a 62-year-old woman to

check for potassium level because she has been on
diuretics for her hypertension for a long time. The
specimen was left in the refrigerator overnight before
delivery to the laboratory the next day. potassium
pseudonyperkalemia

Result: Potassium 6.7 mmol/L (3.5-5.0 mmol/L)

After analysing the result, the biochemist was immediately

on the phone to the doctor in charge.
Why?
resultssupposed
to belowerman
referencerangedueto
diuretics pseudonypercalemia
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