HEMATOLOGY

INTRODUCTION

• Hematology is a discipline concerned with the production,

function, and disorders of blood cells and blood proteins
• Blood is a liquid consisting of plasma (water, electrolytes,
nutrients, waste products, and many soluble proteins) in which
red cells, platelets and a variety of white cells are suspended.
• Blood volume is about 70 mL/kg or about 5L total in an average
size adult.
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Blood Components
Maintain oncotic pressure.
α & β transport lipids &
Clotting Fs i, ii, v fat soluble vit., γ confer

if blood allowed to
coagulate (30min)
& vii removed immune defense.
=Serum /
Blood clotting.

/Thrombocytes
/ Leucocytes

Defined by morphological
differences & functional
Hematocrit:
% of blood

characteristics
that is cells

/ Erythrocytes
Obtained by centrifugation of
whole blood (containing citrate or
heparin anticoagulant) for ≈ 5 min
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INTRODUCTION
• One of the most common tests ordered by all medical specialties is
the complete blood count (CBC) or Full haemogram (FHG).
• The clinical laboratory uses an analyzer that functions both as a
spectrophotomer and a flow cytomer.
• These instruments can determine the concentration (count) of
RBC, platelets and WBC, the hemoglobin (Hgb) concentration,
and the mean size of the red cells (MCV).
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INTRODUCTION
• The flow cytometer can accurately distinguish and count the
various types of WBC, which it reports as a “differential”.
• The hematocrit (Hct), which represents the percentage of blood
volume occupied by red cells, is calculated from the RBC count
and the MCV
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COMPLETE BLOOD COUNT

PARAMETER DESCRIPTION REFEREN CE REFEREN CE
RAN GE (M) RAN GE (F)
Red blood cell count Total number of red blood cells in blood 3.8-5.2 x 1012/L 4.5-6.0 x 1012/L,
( RBC )
Haemoglobin (Hb) Concentration of hemoglobin in blood 13-18 g/dL 12-16 g/dL

Hematocrit (Hct) Volume of “packed” red blood cells. Calculated by 40-52%, 35-47 %
machine (MCV x RBC) (Rough guideline: Hct
should = Hgb x 3).
Mean corpuscular (cell) Mean red blood cell volume 80-100 fL 80-100 fL
volume (MCV)
Mean cell Hgb (MCH) Weight of Hgb in the average red blood cell 26-34 pg 26-34 pg
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l range = Adult: 4.5-11 x 109/L. Child: 5.0-17.0 x 109/L (the exact range differs according

COMPLETE BLOOD COUNT

• White blood cell count (WBC) = total number of leukocytes in blood
• The WBC differential reports each cell type as a percentage of the total WBC.
} Differentials:percentages of each cell type in blood
DIFFERENTIALS NORMAL RANGE NORMAL RANGE
(ABSOLUTE COUNT,
ADULTS) / microL
ADULT CHILDREN
Neutrophils 45-75% 20-55% 1,600 – 6,700
Lymphocytes 20-50% 25-75% 1,300 – 4,200
Monocytes 1-8% 0-10% 110 – 810
Eosinophils 0-6% 0-6% 0 – 575
Basophils 0-1% 0-6% 0 –160
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COMPLETE BLOOD COUNT

• Platelet count (Plt) = total number of platelets in blood
• Normal range of Platelet count = 150-450 x 109/L
• Normal range depends on the platelet count: (Normally, if the
platelet count falls, the body compensates a little by trying to
make bigger platelets.)
• The values of cell counts, hemoglobin, and hematocrit are in part
dependent on the plasma volume.
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EXAMINATION OF BLOOD SMEAR: RBCs

• Look at the red blood cells } Estimate number.

• Look for variation in size (anisocytosis, poikilocytosis). Oval
macrocytes (B12/folate deficiency)
• Microcytes (iron deficiency anemia, thalassemia)
• The size range can often help you narrow down which type of
anemia is present!
• Schistocytes (microangiopathic hemolytic anemia)
• Spherocytes (hemolytic anemia, hereditary spherocytosis)
• Teardrop cells or dacryocytes (myelofibrosis or myelophthisic
processes
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RETICULOCYTES

• Immature red cells that still contain a little ribosomal RNA.

• When you do a supravital stain on a blood smear, the RNA stains
blue, and the cells are called “reticulocytes”.
• When you do a normal Wright-Giemsa stain, the cells look big and
slightly basophilic, and they are called “polychromatophilic cells.”
• Normal reticulocyte count is 0.5-2.5% for adults and 2-6 % for
infants.
• Absolute count: 50-100 x 109/L
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RETICULOCYTES
• Reticulocyte count is a marker of red cell production and helps in
distinguishing hypo and hyper proliferative marrow disorders
• Reticulocyte count is useful in:
Distinguishing the various causes of anemia e.g. For instance in hemolytic
anemia: there is high reticulocyte count to compensate for loss of blood
In an anemic patient, an absolute reticulocyte count of greater than 200,000 is
a good indication that the anemia is not due to inadequate red cell production.
Monitoring of bone marrow response and the return of normal marrow
function following chemotherapy, bone marrow transplant or post treatment
follow up for iron deficiency anemia, vitamin B12 or folate deficiency anemia, or
renal failure
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WBCs: NEUTROPHILS
• The neutrophil (also called polymorphonuclear leukocyte, or
PMN) is usually the most abundant of the WBC in the peripheral
blood.
• It is an ameba-like phagocyte, loaded with a variety of potent
enzymes.
• The first white cell on the scene of inflammation, it lives a
relatively short life of at most 5-7 days after leaving the bone
marrow.
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NEUTROPHILS
• After ingesting and digesting bacteria, the neutrophil self-
destructs from the effects of its released enzymes and oxidants.
• On the peripheral smear, neutrophils are easily identified by their
segmented nucleus and indistinct pink granules in their
cytoplasm.
• They are 15-20 μm in diameter, about three times the size of the
RBC.
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NEUTROPHILS
 Immature neutrophils in which the
nucleus has not completely segmented
are referred to as “bands”; these
increase in number during bacterial
infections because they leave the bone
marrow early.
 About 50% of neutrophils at any given
time are loosely attached to vessel walls
(marginated), but can be released into
circulation with stimuli such as stress or
exercise.
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EOSINOPHILS AND BASOPHILS

 Eosinophils and basophils, the other WBC with large
numbers of prominent granules, normally make up a
small fraction of WBC.
 Increased eosinophils can be seen in association with
asthma or allergic reactions, helminth infections, or in
association with certain malignancies or drug reactions.
 An increase in basophils suggests the presence of a
primary bone marrow proliferative disorder (for
example, chronic myelogenous leukemia).
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LYMPHOCYTES

• Lymphocytes are typically slightly less numerous than neutrophils in

the peripheral blood, except in young children.
• These small (about 10 μm, slightly bigger than RBC), mononuclear
cells have relatively scant, blue-staining cytoplasm and round nuclei
with dense, sometimes clumpy chromatin.
• It is not possible to distinguish T cells from B cells by morphology.
Both can become larger, with increased cytoplasm, upon activation.
• Large granular lymphocytes, a mixture of NK (natural killer) cells and
cytotoxic (CD8+) T cells, make up a special class of lymphocytes.
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LYMPHOCYTES
• Lymphocytes can be quite long-lived, so lymphocyte counts do not
necessarily tell us much about the patient’s current bone marrow
output.
• Increased lymphocytes may indicate lymphoproliferative
neoplasm but also occur in viral and bacterial infections, most
notably in infectious mononucleosis (Epstein-Barr virus) and
whooping cough (Bordetella pertussis).
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LYMPHOCYTES
• These have increased cytoplasm
containing a few purple-red granules.
• They are sometimes referred to as
“atypical lymphocytes”
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MONOCYTES
• The peripheral blood monocyte is a precursor to tissue
macrophages.
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MONOCYTES
• It is sometimes difficult to discriminate
between activated or large granular
lymphocytes and monocytes on the
peripheral smear.
• Compared with the lymphocyte, the
monocyte has more cytoplasm, which tends
to be grey rather than blue, and is larger
than most lymphocytes, with a bean-shaped
or folded nucleus and less dense chromatin.
Monocyte numbers increase in some
myeloproliferative disorders and in some
infections such as tuberculosis.
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THE BONE MARROW AND BLOOD CELL DEVELOPMENT

• All of the cells in peripheral blood
originate in the bone marrow in the adult
human.
• In embryonic life, phases of hematopoiesis
occur in the yolk sac, liver, and spleen,
with the bone marrow becoming dominant
by the time of birth
• Bone marrow biopsy from a normal 40-
year-old man with approximately 60%
cellularity and 40% adipose tissue.
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THE BONE MARROW AND BLOOD CELL DEVELOPMENT

• When it is necessary to evaluate the bone marrow microscopically, a
smear of a liquid aspirate of bone marrow containing marrow cells,
blood, and small spicules of bone is prepared and stained with Wright
stain.
• The biopsy section provides useful information about marrow cellularity
and architecture
• About 50% of marrow cells are neutrophils and their precursors, 25%
erythroid precursors, and the remainder lymphocytes, monocytes,
plasma cells, and others.
• All blood cells originate from a collection of long term hematopoietic
stem cells (LT-HSC)
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HAEMOPOIESIS AND STEM CELLS

• HSC undergo a number of cell divisions and differentiation to

precursor cells with progressively more limited lineage capacities,
eventually giving rise to a large cohort of mature blood cells.
• Except for lymphocytes and perhaps monocytes, these mature
cells are postmitotic (no longer able to divide)
• Flow cytometry-based methods are able to separate these
precursors by virtue of their surface protein phenotype
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