Overview of Clinical

Hematology
Hematology
• Study of the disorder and abnormalities related or associated with
the quality and quantity of the cellular elements of the blood
• Study of the laboratory procedure and techniques being used to
examine the quality and quantity of the cellular elements of the
blood
Overview
• The average human possesses 5 liters of blood
• Blood transports oxygen from lungs to tissues
• Clears tissues
• Transports:
• Moves wastes
• Plasma
• Three categories of blood:
• RBCs
• WBCs
• Platelets
History
• (1657) Athanasius Kircher – described “worms”
• (1674) Anton van Leeuwenhoek – gave an account of RBCs
• (1842) Giulio Bizzozero – described platelets as “petites plaques.”
• (1902) James Homer Wright – Wright Stain
History
• (1658) - discovery of erythrocytes by Swammerdam
• (1846) - PMN distinguished from other leukocytes by Wharton
Jones
• (1879) - first complete classification of leukocytes by Ehrlich
• (1920) - hematology was considered a separate science from
clinical pathology
Basic Hematology Terminology
• a - without • Hypo – decreased
• -blast – youngest / • Iso- equal
nucleated • Macro- large
• -chromic – color
• Mega-very large / huge
• -cyte – cell
• Micro- small
• Dys – abnormal
• -emia – in the blood • Myelo – marrow
• Ferro-iron • Normo – normal
• Hyper- increased • -oid-like
Abbreviation
• FBC – Full Blood Count • MCHC – Mean Cell Hemoglobin
• Fl- Femtoliter Concentration
• Hb – Hemoglobin Concentration • CBC- Complete Blood Count
• Hct- Hematocrit • Pg - Picogram
• MCH – Mean Cell Hemoglobin
• MCV – Mean Cell Volume
Blood Composition
1. FORMED ELEMENTS – Includes erythrocytes, leukocytes,
platelets
2. LIQUID PORTION
A. PLASMA is the liquid portion of unclotted blood

B. SERUM : is the fluid that remains after coagulation has

occurred and clot has formed
Differentiation of Liquid Portion of Serum
and Plasma
General Characteristics of Blood
1. In vivo, blood is in fluid; in vitro it coagulates in 5-10 minutes
2. Thick and viscous 3.5-4.5 to 5 times thicker than water
3. Makes up 6-8 % of the total body component or 60-80ml/kg
4. Total Blood Volume (TBV)
Adult Male : 5-6 L
Adult Female : 4-5 L
Newborn : 250-350 ml
General Characteristics of Blood

5. pH of Blood
• pH is a measure of the acidity or basicity of an aqueous solution
• it is the negative decimal logarithm of hydrogen concentration

Normal pH 7.35 – 7.45 ( average of 7.40 )

venous blood : 7.35
arterial blood : 7.45

** if pH is less than 7.3 ------- A

** if pH is more than 7.5 -------A
pH of the Blood
• pH is maintained by
• Excretion of CO2
• Excretion of H+ or OH
General Characteristics of Blood
6. Blood Specific Gravity – is the ratio of the density of a substance
to the density of a reference substance.
• number of solutes dissolved in the blood
whole blood : 1.045 – 1.066
serum : 1.024 – 1.028
plasma : 1.025- 1.029

Sp. Gravity depends : RBC’s and Plasma proteins

General Characteristics of Blood
7. COLOR
• Arterial Blood : bright red
• Venous blood : dark purplish red
• In pulmonary arteries and veins Hb with O2 : purple
• Hb w/o O2 : blue

8. Osmolality – concentration of solutes dissolved in the blood

• uses OSMOMETER for measurement
• Reference range : 281-297 MILLI-OSMOLES PER KILOGRAM. ( mOsms)
OVERVIEW OF FORMED
ELEMENTS
RED BLOOD CELLS, WHITE BLOOD CELLS AND PLATELETS
Red Blood Cells
• Anucleate, biconcave, discoid cells
• Filled with hemoglobin
• Appear salmon pink
• 7-8um in diameter
• w/ a zone of pallor
Hemoglobin, Hematocrit and Blood Cell
Indices
• Hemoglobin
• Hematocrit
• RBC Indices
• Mean Cell Volume (MCV)

• Mean Cell Hemoglobin (MCH)

• Mean Cell Hemoglobin Concentration (MCHC)

• RBC Distribution Width (RDW)

Reticulocytes
• Polychromatic erythrocytes
• Slightly blue-gray
• Indicate the ability of the bone marrow to increase RBC production
in anemia
• Nucleic acid stains or vital/supravital stains
White Blood Cells
• Protects its host from infection or injury
• Source: Bone Marrow or Lymphoid Tissue
• Leukopenia
• Leukocytosis
• Types: Neutrophils, Eosinophils, Basophils, Lymphocytes and
Monocytes
Neutrophils
• phagocytic cells
• Major purpose is to engulf and destroy microorganisms and foreign
material
• Cytoplasm: pink-lavender staining granules
• Neutrophilia
• Neutropenia
Bands
• Less mature neutrophils with a non-segmented nucleus
• U or S shape
• “left shift”
Eosinophils
• round, bright orange-red cytoplasmic granules
• Eosinophilia
Basophils
• cells with dark purple, irregular cytoplasmic granules that obscure
the nucleus.
• Granules = histamines and proteins
• Basophilia
Lymphocytes
• For host immunity
• Cell-mediated response
• nearly round, are slightly larger than RBCs
• have round featureless nuclei and a thin rim of nongranular
cytoplasm
• Lymphocytosis
• Lymphopenia/Lymphocytopenia
Monocytes
• Immature macrophage
• Identifies and phagocytize foreign particles
• Assist the lymphocytes in mounting an immune response
• have a slightly larger diameter than other WBCs
• blue-gray cytoplasm with fine azure granules
• nucleus that is usually indented or folded
• Monocytosis
Platelets
• maintain blood vessel integrity
• Thrombosis
• major cells that control hemostasis
• 2 to 4 um in diameter
• round or oval, anucleate
• slightly granular
• Thrombocytosis
• Thrombocytopenia
OVERVIEW OF COMPLETE
BLOOD COUNT
CBC
Complete Blood Count
Blood Film Examination
• “wedge-prep”
• WBC Differential Count = 100 WBCs - Percent distribution
THANK YOU!
• REFERENCES:
• Hatton, Christian . (2018). Haematology : lecture notes, 10th ed. New Jersey : John Wiley & Sons. 616.07561 H28 2018,c5

• Turgeon, Mary Louise. (2018). Clinical hematology : theory and procedures, 6th ed.. Philadelphia : Wolters Kluwer.
616.15 T84 2018

• Bain, Barbara J.. (2017). Dacie and Lewis practical haematology, 12th ed. Philadelphia : Elsevier. 616.07561 D11 2017

• Keohane, Elaine M..[et.al]. (2016). Rodak's Hematology : Clinical Principles and Applications, (5th ed.). Singapore: Elsevier.
616.15 R61 2016

• Sadang, Ma. Gina M. (2015). Laboratory manual in hematology. Quezon City : C & E. FR 616.150078 S1 2015

• Antonio Pascua, RMT, MSMT. Hematology 1 Lecture. Our Lady of Fatima University. Valenzuela City.
HEMATOPOIESIS
HEMATOPOIETIC DEVELOPMENT
• Hematopoiesis - is a continuous, regulated process of blood cell
production that includes :
HEMATOPOIETIC DEVELOPMENT
A. EMBRYONIC / FETAL DEVELOPMENT MESOBLASTIC PHASE
• Begins around the 19th day of embryonic development after
fertilization
• Primitive erythrocytes are found in the yolk sac arise from
mesodermal cell
• Cells from the mesoderm migrate to the yolk sac
• Transient yolk sac erythroblasts are important in early embryogenesis
to produce hemoglobin (Gower-1, Gower-2, and Portland)
• Alpha globin chain production begins at this phase
HEMATOPOIETIC DEVELOPMENT
B. HEPATIC PHASE
• The hepatic phase of hematopoiesis begins at 5 to 7 gestational weeks
• Characterized by recognizable clusters of developing erythroblasts,
granulocytes, and monocytes colonizing the fetal liver, thymus, spleen,
placenta
• The developing erythroblasts signal the beginning of definitive
hematopoiesis with a decline in primitive hematopoiesis of the yolk sac
• Hematopoiesis during this phase occurs extravascularly, with the liver
remaining the major site of hematopoiesis during the second trimester of
fetal life
HEMATOPOIETIC DEVELOPMENT
B. HEPATIC PHASE
• The developing spleen, kidney, thymus, and lymph nodes contribute
to the hematopoietic process during this phase.
• Thymus is the first fully developed organ in the fetus
• Kidney and Spleen produce B cells.
• Detectable levels of Hb (F) and Hb A is present.
HEMATOPOIETIC DEVELOPMENT
C. MEDULLARY (MYELOID PHASE)
• Hematopoietic activity, especially myeloid activity, is apparent during
this stage of development, and the myeloid-to-erythroid ratio
gradually approaches 3:1 (adult levels)
• Measurable levels of erythropoietin (EPO), (G-CSF), (GM-CSF),
hemoglobins F and A can be detected.
ADULT HEMATOPOIETIC TISSUE
• In adults, hematopoietic tissue is located in the
•
•
•
•
• Lymphoid development occurs in primary and secondary lymphoid
tissue
POST NATAL DEVELOPMENT
• Hematopoietic tissue is involved in the proliferation and maturation
of blood cells.
BONE MARROW
• Major function of BM is the proliferation and production of blood
cells

TWO MAJOR COMPONENTS

1. RED MARROW
- hematopoietically active marrow consisting of the developing blood
cells and their progenitors
ORGANS INVOLVED IN POST NATAL
DEVELOPMENT: BONE MARROW
TWO MAJOR COMPONENTS
2. YELLOW MARROW
- hematopoietically inactive marrow composed primarily of adipocytes
and (fat cells), with undifferentiated mesenchymal cells and
macrophages
HEMATOPOIETIC MICROENVIRONMENT
• It is responsible for supplying semifluid matrix (stroma) that serves as
an anchor for the developing hematopoietic cells.
Composition of Stroma
1. Endothelial cells
2. Adipocytes
3. Macrophages
4. Osteoblasts
5. Osteoclasts
6. Reticular cells (fibroblasts)
HEMATOPOIETIC MICROENVIRONMENT
Composition of Extracellular Matrix of BM
1. Collagen
2. Fibronectin
3. Thrombospondin
4. Laminin
5. Proteoglycans
ORGANS INVOLVED IN POST NATAL
DEVELOPMENT: LIVER
• liver serves as the major site of blood cell production during the
second trimester of fetal development
Functions:
• Protein synthesis and degradation
• Coagulation factor synthesis carbohydrate and lipid metabolism
• Drug and toxin clearance
• Iron recycling and storage
• Hemoglobin degradation
ORGANS INVOLVED IN POST NATAL
DEVELOPMENT: SPLEEN
• It is located directly beneath the diaphragm behind the fundus of the
stomach in the upper left quadrant of the abdomen
• It is vital but not essential for life

Functions
• indiscriminate filter of the circulating blood
• serves as a storage site for platelets
ORGANS INVOLVED IN POST NATAL
DEVELOPMENT: SPLEEN
3 REGIONS: Two methods for removing
1. White pulp senescent or abnormal RBCs from
the circulation:
2. Red pulp
1. Culling
3. Marginal zone
2. Pitting
ORGANS INVOLVED IN POST NATAL
DEVELOPMENT: LYMPH NODES
• are members of lymphatic system located along the lymphatic
capillaries.

3 Functions:
1. Play a role in the formation of new lymphocytes from germinal
centers.
2. Involved in the processing of specific Ig.
3. Involved in the filtration of particulate matter, debris, and bacteria
entering the lymph node via the lymph
ORGANS INVOLVED IN POST NATAL
DEVELOPMENT: LYMPH NODES
• REGIONS:
• CORTEX
• PARACORTEX
• MEDULLA
ORGANS INVOLVED IN POST NATAL
DEVELOPMENT: THYMUS
• originates from endodermal and mesenchymal tissues.
• is populated initially by lymphocytes from the yolk sac and the liver.
• is an efficient, well-developed organ at birth that consist of two
lobules each measuring 0.5 to 2 cm in diameter. –
STEM CELLS
1. Cells that have extensive proliferative capacity
2. HSC are BMC that are capable of producing all types of blood cells
3. They differentiate into one or another type of committed stem cells
HEMATOPOIETIC GROWTH FACTORS
1. CSF : Colony Stimulating Factors
A. GM-CSF
- a pan myeloid growth factor that stimulates granulo, mono,
megakaryocyte and eosinophil progenitors
- Source : fibroblast, T cells and endothelial cells
B. G-CSF
- stimulates granulocytes production and functional activation
- Source : monocytes and fibroblast
HEMATOPOIETIC GROWTH FACTORS
C. M-CSF
- stimulates monocytes and macrophages production activity
- Source : monocytes, fibroblast, endothelial cells.

D. Meg-CSF
- Specific to the megakaryocyte lineage
– source : monocytes, fibroblast , megakaryocytes
HEMATOPOIETIC GROWTH FACTORS
2. Erythropoietin (EPO )
- Stimulates proliferation , growth and differentiation of erythroid
precursors and may have minor effects on megakaryocytes.
- Target cells are pronormoblast and CFU-Erythroid cells
- Source: Kidney

3. Thrombopoietin
- regulates production platelets.
CONTROL OF HEMATOPOIESIS
The entry of mature blood cells into the intravascular space relies
upon:
1. Multiplication of developing cells
2. Gradual maturation
3. Orderly release of cell from bone marrow
3 POSSIBLE ACTIONS OF HSCs
1. Self-renewal
2. Differentiation
3. Apoptosis
CYTOKINES
• is a group of specific glycoproteins called growth factors that
regulates the proliferation differentiation, and maturation of
hematopoietic precursor cells

– includes:
• Interluekins (Ils)
• Lymphokines
• Monokines
• Interferons
• Colony Stimulating Factors (CSFs)
• Chemokines
CYTOKINES
POSITIVE INFLUENCE NEGATIVE INFLUENCE
• 1. IL-1 1. Transforming Growth Factor-β
• 2. IL-3 2. Tumor Necrosis Factor-α
• 3. IL-6 3. Interferons
• 4. IL-9
• 5. IL11
• 6. GM-CSF
• 7. Kit Ligand
THANK YOU!
• REFERENCES:
• Hatton, Christian . (2018). Haematology : lecture notes, 10th ed. New Jersey : John Wiley & Sons. 616.07561 H28 2018,c5

• Turgeon, Mary Louise. (2018). Clinical hematology : theory and procedures, 6th ed.. Philadelphia : Wolters Kluwer.
616.15 T84 2018

• Bain, Barbara J.. (2017). Dacie and Lewis practical haematology, 12th ed. Philadelphia : Elsevier. 616.07561 D11 2017

• Keohane, Elaine M..[et.al]. (2016). Rodak's Hematology : Clinical Principles and Applications, (5th ed.). Singapore: Elsevier. 616.15
R61 2016

• Sadang, Ma. Gina M. (2015). Laboratory manual in hematology. Quezon City : C & E. FR 616.150078 S1 2015

• Antonio Pascua, RMT, MSMT. Hematology 1 Lecture. Our Lady of Fatima University. Valenzuela City.
ERYTHROCYTES
ERYTHROCYTES
• Mature RBCs size 7-8 um ave in diameter
• has no nucleus nor organelles
• Exists in blood circulation for 120 days
• Limited activity to metabolize fatty acids and amino acids
• Metabolic processes are maintained through different
• metabolic pathways to produce energy
RBC MEMBRANE CHARACTERISTICS
• RETICULOCYTE MEMBRANE
• Possesses a significant amount of tubulin and actin
• Changes:
• Increase in shear resistance
• Loss of surface area (about20%)
• Acquisition of a biconcave shape
• Loss of cytoplasmic organelles
• Undergoes active endocytosis and exocytosis which does
• not occur in mature RBCs
RBC MEMBRANE CHARACTERISTICS
• MATURE RED BLOOD CELL MEMBRANE

• Soft and pliable

• Biconcave shape
• Consists of a membrane skeleton protein lattice and lipid
bilayer
RBC MEMBRANE CHARACTERISTICS
• MATURE RED BLOOD CELL MEMBRANE
• Deformable and tolerant against mechanical stress and various
pH and salt concentrations in vivo and in vitro
• Cell shape changes reversibly
STRUCTURE OF AN RBC MEMBRANE
• MEMBRANE LIPIDS • Glycophorin
• Aquaporin
Outer layer
• Phosphatidyl choline
• Sphingomyelin PERIPHERAL PROTEIN
• Spectrin
Inner layer
• Actin
• Phosphatidyl ethanolamine
• Protein 4.1
• Phosphatidyl serine
• Pallidin (band 4.2)
• Ankyrin
• MEMBRANE PROTEINS • Adducin
Integral Protein • Tropomycin
• Band 3 (anionexchanger protein) • Tropomodulin
BAND 3
• Anion transport
• Exchanges bicarbonate or chloride
• Linkage of lipid bilayer to the underlying membrane skeleton
GLYCOPHORIN
• Imparts a negative charge to the cell
• Glycophorin A -
• Glycophorin C, Glycophorin A –
AQUAPORIN 1
• Selective pores for water transport
RED CELL MEMBRANE SKELETON
• Hexagonal lattice with 6 spectrin moleucles
• Each are linked to multiple spectrin tetramers
• Composition:
• Spectrin
• Actin 4.1
• Ankyrin
ACTIN
• Short, uniform filaments
• Length is modulated by tropomyosin/tropomodulin
OTHER PERIPHERAL PROTEINS
• PROTEIN 4.1
• Stabilizes actin-spectrin interactions

• ADDUCIN
• Also stabilzes interaction of spectrin with actin

• ANKYRIN
• Interacts with band 3 and spectrin to achieve linkage between
bilayer and skeleton
RED CELL MECHANICS
• INFLUENCED BY:
• CELL SHAPE
• CYTOPLASMIC VISCOSITY
• MEMBRANE DEFORMABILITY AND STABILITY
CELL SHAPE
• Facilitates deformation while maintaining constant surface
area
• Progressive loss of intracellular and membrane
components results in biconcave shape and improved
deformability.
• SA/V Ratio alterations result in more spherical shape with
less redundant surface area
MEMBRANE DEFORMABILITY/STABILITY
• During pressure upon RBC
• During extreme or sustained pressure
• Deformability can be reduced by increases in associations
between skeletal proteins or between skeletal and
integral proteins.
CYTOPLASMIC CHARACTERISTICS
• Contents:
• Potassium ions
• Sodium ions
• Glucose
• Intermediate products of glycolysis
• Enzymes
METABOLIC ACTIVITIES
ENERGY METABOLISM IN THE
ERYTHROCYTE
EMBDEN-MEYERHOF PATHWAY
• Glucose undergoes glycolysis to
form ATPs
• Maintains pyridine nucleotides in
reduced state to permit their
function in oxidation-reduction
reactions within the cell
• Deficiencies to production of ATP
can be exhibited by:
• Premature cell death due to
inherited defects in glycolysis
• Loss of viability during the storage of
blood for transfusion
HEXOSE MONOPHOSPHATE SHUNT
• Oxidative catabolism of glucose
with reduction of NADP to NADPH
• If the pathway is defective:
• Amount of reduced glutathione
becomes insufficient to neutralize
oxidants
METHEMOGLOBIN REDUCTASE
PATHWAY
• Prevents the oxidation of heme
iron
• Requires the reducing action of
NADH and the enzyme
methemoglobin reductase
LEUBERING-RAPOPORT PATHWAY
• This mechanism is low in energy
consumption
• Capable of regulating oxygen
transport even with hypoxia and
acid-base disorders
• Permits the accumulation of 2,3
DPG
• Increased deoxyhemoglobin
results to binding of 2,3 DPG
METABOLIC PATHWAYS IN THE
ERYTHROCYTE
ERYTHROKINETICS
• Term describing the dynamics of RBC production and
destruction

• Erythron - - name given to the collection of all stages of

erythrocytes throughout the body, developing precursor
in the bone marrow and the circulating RBC in the
peripheral blood.
ERYTHROPOIETIN
• SPECIFIC ACTION:
• induces committed progenitor cells in the bone marrow to
differentiate and proliferation into pronormoblast
• . Shortens the generation time of pronormoblast
• Promotes the early release of reticulocytes to the peripheral
blood.
MECHANISM OF RED CELL
DESTRUCTION
• FRAGMENTATION
- Loss of a portion of the erythrocytes membrane
• OSMOTIC LYSIS
- Passing of water into the red cell
• ERYTHROPHAGOCYTOSIS
-Ingestion of the whole RBCs by a circulating monocytes or
neutrophils or by macrophages
• COMPLEMENT INDUCED CYTOLYSIS
- Complement attach itself to the cells and induce lysis
• HEMOGLOBIN DENATURATION
- When Hgb is exposed to oxidant stress
2 TYPES OF DESTRUCTION
• INTRAVASCULAR
- lysis of erythrocytes which occurs within the circulation through
the classic pathway
2 TYPES OF DESTRUCTION
• INTRAVASCULAR
Causes:
ABO mismatched blood transfusion
Cold agglutinin disease
Paroxysmal cold hemoglobinuria
Burns
Snake bites
Bacterial- C. perfringens
Parasitic infections- P. malaria
Mechanical heart valves
Paroxysmal nocturnal hemoglobinuria
2 TYPES OF DESTRUCTION
• EXTRAVASCULAR
- lysis of erythrocytes outside the
circulation
- Little or no hemoglobin escapes into
the circulation
- Decreased haptoglobin
- Normal Plasma Hemoglobin
2 TYPES OF DESTRUCTION
• EXTRAVASCULAR
Causes:
Bacterial/Viral infections
Drug induced
Autoimmune
Microangiopathy
Hemoglobinopathies
Membrane defects
Metabolic defects-G6PD
THANK YOU!
• REFERENCES:
• Hatton, Christian . (2018). Haematology : lecture notes, 10th ed. New Jersey : John Wiley & Sons. 616.07561
H28 2018,c5

• Turgeon, Mary Louise. (2018). Clinical hematology : theory and procedures, 6th ed.. Philadelphia : Wolters
Kluwer. 616.15 T84 2018

• Bain, Barbara J.. (2017). Dacie and Lewis practical haematology, 12th ed. Philadelphia : Elsevier. 616.07561
D11 2017

• Keohane, Elaine M..[et.al]. (2016). Rodak's Hematology : Clinical Principles and Applications, (5th ed.).
Singapore: Elsevier. 616.15 R61 2016

• Sadang, Ma. Gina M. (2015). Laboratory manual in hematology. Quezon City : C & E. FR 616.150078 S1 2015

• Antonio Pascua, RMT, MSMT. Hematology 1 Lecture. Our Lady of Fatima University. Valenzuela City.
HEMATOPOIETIC SYSTEM
LINEAGE SPECIFIC HEMATOPOIESIS
THE PRINCIPLE OF NORMAL BLOOD CELL
MATURATION
I. CYTOPLASMIC CHANGES
II. CYTOPLASMIC GRANULES
III. NUCLEAR CHANGES
IV. REDUCTION IN CELL SIZE
THE PRINCIPLE OF ABNORMAL CELL
MATURATION
I. ABNORMAL CYTOPLASMIC DIFFERENTIATION
II. ABNORMAL NUCLEAR MATURATION
III. ABNORMAL SIZE
GENERAL CHARACTERISTIC OF A BLAST
1. Size : large cell with high N: C ratio
2. Cytoplasm : very dark blue and small in
amount in comparison to the size of the
nucleus. No granular is present.
3. Nucleus : large in size as compared to the
size of cytoplasm.
Chromatin which is reddish purple and
indicates predominance of DNA.
HEMATOPOIETIC SYSTEM
• Erythropoiesis
• Granulopoiesis
• Monopoiesis
• Lymphoiesis
• Megakaryopoiesis
ERYTHROPOIESIS
ERYTHROPOIESIS
• is a process by which erythroid precursor cells differentiates to
become mature.
ERYTHROPOIESIS
• PRONORMOBLAST (RUBRIBLAST)
Nucleus
• takes up much of the cell (N:C ratio of 8:1)
• round to oval
• contains one or two nucleoli.
• the purple red chromatin is open and contains
few, fine clumps.
Cytoplasm
• dark blue
ERYTHROPOIESIS
• PRONORMOBLAST (RUBRIBLAST)
Location
• present only in the bone marrow in healthy
states
Cellular Activity
• begins to accumulate the components
necessary for hemoglobin production.
• The proteins and enzymes necessary for iron
uptake and protoporphyrin synthesis are
produced.
ERYTHROPOIESIS
• BASOPHILIC NORMOBLAST (PRORUBRICYTE)
Nucleus
• The chromatin begins to condense
• N:C ratio decreases to about 6:1.
• The chromatin stains deep purple-red.
• Nucleoli may be present early in the stage but
disappear later
Cytoplasm
• When stained it is deeper, richer blue in color than
in the pronormoblast

Location
• present only in the bone marrow in healthy states
ERYTHROPOIESIS
• BASOPHILIC NORMOBLAST
(PRORUBRICYTE)
Cellular Activity
• Detectable hemoglobin synthesis occurs,
but the many cytoplasmic organelles,
including ribosomes and a substantial
amount of messenger ribonucleic acid,
completely mask the minute amount of
hemoglobin pigmentation
ERYTHROPOIESIS
• POLYCHROMATIC (POLYCHROMATOPHILIC)
NORMOBLAST (RUBRICYTE)
Division
• This is the last stage in which the cell is capable of
undergoing mitosis

Location
• present only in the bone marrow in healthy states.

Cellular Activity
• Hemoglobin synthesis increases, and the
accumulation begins to be visible in the color of the
cytoplasm.
ERYTHROPOIESIS
• ORTHOCHROMIC NORMOBLAST (METARUBRICYTE)
Nucleus
• completely condensed
• N:C ratio is low or approximately 1:2.
Cytoplasm
• The increase in the salmon-pink color of the cytoplasm
reflects nearly complete hemoglobin production
Division
• The orthochromic normoblast is not capable of
division due to the condensation of the chromatin.

Location
• present only in the bone marrow in healthy states
ERYTHROPOIESIS
• RETICULOCYTE
• No nucleus but has mitochondria and
ribosomes
• Last stage to synthesize hemoglobin
• Last stage in bone marrow before release to
the blood
Location
• resides in the bone marrow for 1 day or
longer and then moves into the peripheral
blood for about 1 day before reaching
maturity
ERYTHROPOIESIS
• RETICULOCYTE

Reference Range : 0.5-1.5 %

–Newborn : 2-6 %
• Supravital stain is used
• Also known as :
Polychromatophilic erythrocytes
Diffusely basophilic erythrocytes
Polychromatophilic macrocytes
ERYTHROPOIESIS
• ERYTHROCYTE
Nucleus
• No nucleus is present in mature RBCs.
Cytoplasm.
• The mature circulating erythrocyte is a
biconcave disc measuring 7 to 8 um in
diameter, with a thickness of about 1.5 to
2.5 mm
• On a stained blood film, it appears as
• a salmon pink-staining cell with a central
pale area.
• The central pallor is about one third the
diameter of the cell.
NOMENCLATURE FOR ERYTHROID
PRECURSORS
ERYTHROCYTE MATURATION SEQUENCE
ERYTHROCYTE MATURATION SEQUENCE
GRANULOPOIESIS
NEUTROPHIL DEVELOPMENT
• MYELOBLASTS
• make up 0% to 3% of the nucleated cells in the bone marrow
and measure 14 to 20 um in diameter

Cytoplasm
• Clear blue, more heavily colored at its border.
• It is non-granular or may have a few azurophilic granules,
depending on the stage of development.

Nucleus:
• Large, round or oval and occupies about four fifths of the
total cell area.
• It has a very fine chromatin meshwork.
• Round / oval nucleus with fine reddish purple staining

Nucleoli : 2 -5
NEUTROPHIL DEVELOPMENT
• PROMYELOCYTES
• comprise 1% to 5% of the nucleated cells in the bone
marrow
• 16 to 25 um in diameter.
Nucleus
• round to oval and is often eccentric
• occupies half or more of the cell
Cytoplasm
• evenly basophilic
• full of primary granules

Nucleoli – 1-3
NEUTROPHIL DEVELOPMENT
Primary (Azurophilic) Granules
Contain:
• Myeloperoxidase
• Acid b-glycerophosphatase
• Cathepsins
• Defensins
• Elastase
• Proteinase-3
NEUTROPHIL DEVELOPMENT
• NEUTROPHIL MYELOCYTES
- make up 6% to 17% of the nucleated cells in
the bone marrow and are the final stage in
which cell division (mitosis) occurs
- During this stage, the production of primary
granules ceases, and the cell begins to
manufacture secondary neutrophil granules.
NEUTROPHIL DEVELOPMENT
• NEUTROPHIL METAMYELOCYTES
- constitute 3% to 20% of nucleated marrow
cells.
- the cells are no longer capable of division,
and the major morphologic change is in the
shape of the nucleus
NUCLEUS
• indented
• chromatin is increasingly clumped.
• Nucleoli are absent
NEUTROPHIL DEVELOPMENT
• NEUTROPHIL BANDS
- make up 9% to 32% of nucleated marrowcells and
0% to 5% of the nucleated peripheral blood cells.
- All evidence of RNAis absent
- tertiary granules continue to be formed during this
stage.
- Secretory granules may begin to be formed during
this stage
NUCLEUS
• highly clumped
• nuclear indentation that began in the metamyelocyte
stage now exceeds one half the diameter of the nucleus
NEUTROPHIL DEVELOPMENT
• SEGMENTED NEUTROPHILS
- make up 7% to 30% of nucleated cells in the
bone marrow
- Secretory granules continue to be formed
during this stage
EOSINOPHIL DEVELOPMENT
EOSINOPHIL DEVELOPMENT
• EOSINOPHILIC PROMYELOCYTES
- can be identified cytochemically due to the
presence of Charcot Leyden crystal protein in
their primary granules
EOSINOPHIL DEVELOPMENT
• EOSINOPHIL MYELOCYTES
are characterized by the presence of large
pale, reddish orange secondary granules,
along with azure granules in blue cytoplasm
EOSINOPHIL DEVELOPMENT
• EOSINOPHIL METAMYELOCYTES AND
BANDS
- resemble their neutrophil counterparts with
respect to their nuclear shape
- Secondary granules increase in number, and
a third type of granule is generated
EOSINOPHIL DEVELOPMENT
• MATURE EOSINOPHILS
Nucleus
• usually display a bilobed nucleus.
Cytoplasm
• contains characteristic refractile, orange-red
secondary granules
EOSINOPHIL GRANULES
• PRIMARY GRANULES
Contain: • Cathepsin D (core and matrix)
• Charcot-Leyden crystal protein
• Interleukins 2, 4, and 5 (core)
• SECONDARY GRANULES • Interleukin-6 (matrix)
Contain:
• Major basic protein (core) • Granulocyte-macrophage
• Eosinophil cationic protein (matrix)
colonystimulating factor (core)
• Eosinophil-derived neurotoxin (matrix)
• Eosinophil peroxidase (matrix)
• Lysozyme (matrix)
• Catalase (core and matrix)
• b-Glucuronidase (core and matrix)
EOSINOPHIL GRANULES
• SMALL LYSOSOMAL GRANULES LIPID BODIES
• Acid phosphatase • Cyclooxygenase
• Arylsulfatase B • 5-Lipoxygenase
• 15-Lipoxygenase
• Catalase
• Leukotriene C4 synthase
• Cytochrome b558
• Eosinophil peroxidase
• Elastase • Esterase
• Eosinophil cationic protein
STORAGE VESICLES
• Carry proteins from secondary
granules to be released into the
extracellular
BASOPHIL DEVELOPMENT
BASOPHIL DEVELOPMENT
• IMMATURE BASOPHILS
Nuclei
• have round to lobulated nuclei with only
slightly condensed chromatin.
• Nucleoli major may not be apparent.
Cytoplasm
• The cytoplasm is blue and contains large blue-
black secondary granules
• Primary azure granules may or may not be
seen.
BASOPHIL DEVELOPMENT
• MATURE BASOPHILS
Nucleus
• contain a lobulated nucleus that is often
obscured by its granules
• The chromatin pattern, if visible, is clumped.
• Actual nuclear segmentation with visible
filaments occurs rarely
Cytoplasm
• colorless and contains large numbers of the
characteristic large blue-black granules.
MONOCYTE DEVELOPMENT
• MONOBLAST
Size : 12-20 um
Cytoplasm : moderately basophilic to blue
or gray ; non granular
Chromatin is fine, lacey
Nucleoli: 1-2
N/C ratio : 4:1 to 3:1
MONOCYTE DEVELOPMENT
• PROMONOCYTES
- are 12 to 18 um in diameter
Nucleus
• slightly indented or folded.
• Chromatin pattern is delicate
• at least one nucleolus is apparent.
Cytoplasm
• blue and contains scattered azure granules
that are fewer and smaller than those seen
in promyelocytes
MONOCYTE DEVELOPMENT
• MONOCYTES
- appear to be larger than neutrophils
(diameter of 15 to 20 um)
Nucleus
• may be round, oval, or kidney shaped, but more
frequently is deeply indented or folded on itself
• Chromatin pattern is looser than in the other
leukocytes and has sometimes been described
as lacelike or stringy
Nucleoli are generally not seen with the light
microscope;
LYMPHOCYTE DEVELOPMENT
LYMPHOCYTE DEVELOPMENT
• LYMPHOBLAST
- 10-18 um
Cytoplasm
• no granules present .
• Mod to dark blue
Nucleus
• Chromatin Pattern is coarse
• round oval in shape,
• it contains one or two nucleoli

N/C ratio is 4:1

LYMPHOCYTE DEVELOPMENT
• PROLYMPHOCYTE
- Size: same with lymphoblast or
smaller
Cytoplasm
• moderate to dark blue and
usually non granular
Nucleus
• round oval chromatin pattern is
more clumped

NC ratio : 4:1
LYMPHOCYTE DEVELOPMENT
• MATURE LYMPHOCYTE

3 Categories :
Small
Medium
Large
LYMPHOCYTE DEVELOPMENT
• MATURE LYMPHOCYTE: SMALL
- Size : 8 -10 um in diameter
Cytoplasm
• usually forms a thin rim around the
nucleus.
• Moderate blue to dark blue

Nucleus
• round oval in shape slightly
indented.

No nucleoli are visible

LYMPHOCYTE DEVELOPMENT
• MATURE LYMPHOCYTE: MEDIUM
- Size: 10-12 um in diameter
Cytoplasm
• more abundant pale to moderate blue.

Nucleus
• round or oval in shape may be slightly
indented, no nucleolus is visible
LYMPHOCYTE DEVELOPMENT
• MATURE LYMPHOCYTE: LARGE
- Size: 12-16 um in diameter
Cytoplasm
• abundant, clear, very pale blue may not
contain few non specific azurophilic
granules.

Nucleus
• round oval in shape and may be slightly
indented.

No nuleolus is visible
MEGAKARYOPOIESIS
MEGAKARYOPOIESIS
• MEGAKARYOBLAST (STAGE I)
- Size: 14-18 um in diameter
Cytoplasm
• varying shades of blue, may have
small blunt pseudopods
• narrow band around the nucleus
• as the cell matures the amount of
cytoplasm increases
Nucleus
• round oval kidney shaped
• N/C is 3:1
MEGAKARYOPOIESIS
• PROMEGAKARYOBLAST (STAGE II)
- S: 15-40 um in diameter

Cytoplasm
• more abundant than in previous stage;
granules begin to form in the golgi
region

Nucleus
• chromatin becomes more coarse
• multiple nucleoli are visible
• N/C ratio : 4:1 to 7:1
MEGAKARYOPOIESIS
• MEGAKARYOCYTE ( STAGE III)
- Size: 30-50 um in diameter

Cytoplasm
• contains coarse clumps of granules
aggregating into little bundles which bud
off from the periphery

Nucleus
• multiple nuclei are present or multilobed

N/C ratio less than 1:4

MEGAKARYOPOIESIS
• PLATELETS
- S: 2-4 um in diameter

Cytoplasm
• light blue to purple
THANK YOU!
• REFERENCES:
• Hatton, Christian . (2018). Haematology : lecture notes, 10th ed. New Jersey : John Wiley & Sons. 616.07561 H28 2018,c5

• Turgeon, Mary Louise. (2018). Clinical hematology : theory and procedures, 6th ed.. Philadelphia : Wolters Kluwer.
616.15 T84 2018

• Bain, Barbara J.. (2017). Dacie and Lewis practical haematology, 12th ed. Philadelphia : Elsevier. 616.07561 D11 2017

• Keohane, Elaine M..[et.al]. (2016). Rodak's Hematology : Clinical Principles and Applications, (5th ed.). Singapore: Elsevier.
616.15 R61 2016

• Sadang, Ma. Gina M. (2015). Laboratory manual in hematology. Quezon City : C & E. FR 616.150078 S1 2015

• Antonio Pascua, RMT, MSMT. Hematology 1 Lecture. Our Lady of Fatima University. Valenzuela City.