This document provides an overview of hematology and blood components. It defines hematology as the study of blood and describes the main components of blood including plasma, red blood cells, white blood cells, and platelets. It then discusses key hematological parameters and how blood analysis can be used for disease diagnosis by identifying deviations from normal physiological values.

1. HAEMATOLOGY
Gabriel Mbassa

2. Definition and scope
Haematology is the
study of blood

3. COURSE DESCRIPTION
• BLS 110 Hematology: 1 credit
(20 Lect., 20 Pract.)
• Pre-requisite: None
• Objective: To impart on
students the basic knowledge
on haematology

4. Course contents:
• Theory; Introduction to haematology.
Haematopoiesis.
• Blood cells. Haemoglobin and
haemoglobinopathies, anaemias,
• Blood grouping systems, blood
coagulation, blood transfusion
practices and blood transfusion
reactions.

5. • Practical: Blood group typing, Cell
counting, Determination of
Heamatological parameters

6. • Blood
• Specialized fluid tissue circulating
through vascular channels
• Carries compounds to all cells
• Receives waste products of
metabolism for transport to organs
of excretion

7. • Maintains haemostasis & defense
of body.
• Blood cells form in bone marrow
• Blood makes 7-8% of body weight
• Extracellular substances (plasma)
make 45-65% of blood
• Blood cells (called formed
elements) form 35-55%.

8. Haematology basis for
diagnosis
• Numerical values of all the
parameters in the blood are
kept within narrow
physiological limits in healthy
animals

9. • Blood parameter levels may are
altered in disease, increase above
or decrease below physiological
values
• Deviations from physiological
values of healthy animals allow
diagnosis of presence of disease

10. • Each disease changes blood values
in its own way, diagnosis is done by
blood analysis
• Common diseases detected by
blood analysis; blood parasites
(intracellular & extracellular),
helminthes, diseases affecting liver,
kidney, other organs, using
functional tests

11. Functions of blood
• Supply nutrients to cells & tissues
• Supply of oxygen to & removal of
CO2 from tissues & cells
• Remove breakdown products,
taking them to excretory organs

12. • Transport surplus metabolites to
storage organs
• Regulate water & electrolyte
metabolism
• Regulate body temperature
• Contain body's defenses against
foreign substances & pathogenic
organisms

13. • Blood is pumped from central
pumping organ Heart, in birds &
mammals divided into two halves
(double heart);
• Arterial part (left), carrying
oxygenated blood
• Venous (right) part with blood rich
in carbon dioxide

14. • Arterial & venous halves of
heart each consist of atrium &
ventricle, supplying separated
vascular systems;
• 1. Systemic (large) circulation
• 2. Pulmonary or small
circulation

15. Blood Composition
• Blood consists of;
Plasma, viscous fluid that
coagulates
Blood cells; Platelets (non
cellular), erythrocytes,
leukocytes

16. Plasma; aqueous solution of
• Proteins (albumin, fibrinogen,
globulins) & blood sugar
• Inorganic substances, Na, K, Ca,
Mg ions, which maintain
chemico-physical properties of
blood. Bicarbonate & phosphate
ions buffer extreme high or low
pH (Co2, lactic acid).

17. • Final pH of blood is 7.4
• Lipids occur in fine suspension
• Plasma carries nutrients,
hormones, enzymes, antigens,
antibodies, & antitoxins for
neutralization of foreign
protein & bacterial toxins

18. • Blood clots as it leaves vessels, a
vital protective mechanism.
• Coagulation depends on ability of
fluid fibrinogen to interact with
thrombin & transform into a
delicate elastic network of fibrin

19. • Coagulation is initiated by
breakdown of thrombocytes,
involves 30 factors.
• Blood in container clots
unless anticoagulant is
added

20. • Coagulation; 1-2 min in birds,
10-15 pig, 10-20 in equine
• After clot cells are trapped in
fibrin, leave clear fluid called
serum, which contains
antibodies

21. • If blood is mixed with an
anticoagulant & left
undisturbed blood cells will
settle down

22. • The duration of the process is
called sedimentation rate (SR)
• Erythrocyte SR (ESR) is
diagnostic in equine, with large
red blood cells (RBC)

23. • That means mean corpuscular
volume (MCV) of > 30x1015 l
(30 femtolitres or 30 fl)
• ESR not diagnostic in small
ruminants (MCV < 30)
• The clear fluid remaining is
plasma

24. 2. BLOOD CELLS
• Blood cells are also called formed
elements of blood.
• Blood is fluid connective tissue having
cells (35-55% of blood) and
extracellular fluid intravascular (blood
plasma) (45-65% of blood)
• Total amount of blood in man is 5 L
(7‑8% bwt)

25. • Blood cells are;
• 1. Red blood cells or
erythrocytes
• 2. White blood cells or
leukocytes
• 3. Platelets or thrombocytes.

26. • Erythrocytes & platelets
functions in blood stream
• Leukocytes function temporarily in
blood & leave by walls of capillaries
& venules to settle in connective
tissues & lymphoid organs.

27. • Some leukocytes return to
the blood stream, but most
end their lives in tissues

28. • Platelets are not true cells in
higher vertebrates. They lack
nucleus as erythrocytes
• In fish, amphibians, reptiles &
birds platelets are nucleated
cells, called thrombocytes

29. • Leukocytes are eukaryotic cells
containing nucleus.
• Five types of leukocytes occur,
classified on basis of presence
or absence of intracytoplasmic
granules as granular & non-
granular (or agranular)
leukocytes

30. • Granular leukocytes further
subdivided according to
stainability of their cytoplasmic
granules into
• Neutrophilic
• Eosinophilic, &
• basophilic granulocytes

31. • Non-granular leukocytes comprise
monocytes and lymphocytes
• Leukocytes divided are also divided
on basis of shape of nucleus as
• 1. mononuclear (leukocyte with
non-lobed nucleus)
• 2. polymorphonuclear (leukocyte
with multi-lobed nucleus)
leukocytes

32. LYMPHOCYTES
• Lymphocytes play role in
reception of foreign molecules
processing & execution of
immune responses.
• Lymphocytes are heterogeneus
morphologically and functionally.

33. Morphological classification
• Classified as small (6-9μm) medium
sized and large (9–15 μm)
• Cell size, cytoplasmic basophilia,
nuclear chromatin indicate age and
maturity
• Immature lymphocytes are large,
basophilic and have smooth chromatin

34. • Mature lymphocytes have decreased
• 1) Nuclear size
• 2) Cytoplasmic basophilia
• 3) DNA content
• 4) Histone dye-binding capability
• Have increased
• 1) Chromatin clumping
• 2) Nucleus – to-cytoplasm ratio

35. • When small lymphocytes are
activated these properties
reverse and the cells undergo
multiplication
(lymphoblastogenesis) in vivo
and in vitro.

36. Functional classification
• Lymphocytes are grouped on basis of
immune responses.
• Lymphocytes involved in cell
mediated immunity and
immunoregulatory functions are
called thymus derived, thymus –
dependent, thymus processed or T-
lymphocytes (T-cells).

37. • Lymphocytes concerned with
formation of humoral
antibodies are thymus
independent, bursa – derived
(in birds), bone marrow
derived (mammals) or B-
lymphocytes (B cells).

38. • Peripheral blood lymphocytes are 80 –
95% T and B types
• Lymphocytes that make 5-20% of
peripheral blood lymphocytes are non-
T, non- B or null cells. These have
different markers and functions.

39. • Many subpopulations of lymphocytes
are found in peripheral blood because
of production, release and
recirculation of lymphocytes at
different stages of maturation &
immunocompetence.
• Different lymphoid organs have
different types of lymphocytes

40. • stained on blood smears by any
Romanowsky stains
• Lymphocytes are round to oval
• Cytoplasm is scanty to moderate
• Cytoplasm varies in basophilia
• Nucleus is spherical to ovoid
• Nuclear membrane distinct

41. • Chromatin is heavily clumped.
• Nucleus may be indented
• Lymphocytes show slow amoeboid
movements to being motile
• Nucleus presents patchy, dense
clumps of heterochromatin (small
lymphocytes).
• Moderate dispersed chromatin
(medium and large sized
lymphocytes).

42. • Fairly dispersed chromatin
(lymphoblasts)
• Distinct nuclear membrane,
and nuclear pores
• Not frequent nucleoli
• Some nuclear pockets occur in
leukemia

43. • Thoracic lymph duct show nuclear
body proximal to nucleolus
• Nucleolus has electron dense area
surrounded by fibrillar material
• Small lymphocytes have very scanty
cytoplasm

44. • Numerous free cytoplasmic
ribosomes
• Few mitochondria
• Small Golgi complex
• Little rough endoplasmic
reticulum

45. • Medium sized lymphocytes have some
extra cytoplasm (moderate to
abundant).
• Various organelles are prominent
• Centrioles are more conspicuous
• There are few pinocytotic vesicles
• Glycogen granules are seen in some
cells

46. • There are also microfilaments and
microtubules (uropod lymphocytes)
• Lysosomes are rare.
• Some granules of variable size appear
(5-15% of T-lymphocytes) called
azurophilic granules

47. • Azurophilic granules are peroxidese
negative. Some cells present large
spherical refractile structure, called
Gall body.
• Some cells show aggregates of
reddish – purple granules (Kurbff
body).
• Ribosome and polyribosome population
vary with stage of cellular maturity
and activity

48. • Free ribosomes are not
engage in protein synthesis.
• Polyribosomes are engaged in
protein synthesis antibody
formation in B-cells and
plasma cells.
• Lymphokine production in T
cells

49. • On scanning electron
microscopy
• Lymphocytes are globular
• Surfaces are smooth or have
some villi
• Surfaces may have fingerlike
projections

50. Biochemistry
• many hydrolytic and oxidative
enzyme in lymphocytes.
• Pre-B and Pre-T cells have terminal
deoxyribonucleotidase (Tdt)
• B-lymphocytes, plasmablasts and
plasma cells stain for 5’ nucleotidase.

51. • T cells and activated B cells
contain lysosomal enzymes;
acid phosphatase, β-
glucoronidase, and acid α-
naphythyl acetate esterase.
These enzymes core not
present in mature B cells

52. • Lymphocytes lack cytoplasmic alkaline
phosphatase, peroxidase, sudan black
reactivity and lysozyme.
• Lymphocytes derive energy by glucose
metabolism (glycolytic pathway)

53. • The pentose phosphate
pathway is not common, but
stimulated under aerobic
conditions.
• Lymphocytes synthesize
protein, glycogen, and fatty
acids.

54. Characterization of B and T
lymphocytes
• Detection of surface antigens by
monoclonal antibodies & flow cytometry.
• Demonstration of surface or cytoplasmic
immunoglobulins by fluorescence and
autoradiography
• Detection of immunologic and non
immunologic receptors by using
heterologous erythrocytes (E-rosettes)

55. • Detection of receptors for Fc of Ig or
complement components (C3b and C3d
receptors) by rosettle formation with
antibody coated eryrothcytes (EA-
rosetes) or antibody-complement –
coated erythrocytes (EAC-rosettles)
• Immunofluorescence & autoradiography
• Bacterial adherence to lymphocytes (in
conjuction with fluoresent antibody
technique).
• Helix prometia antigen chracterizes
bovine and equine T-lymphocytes
• Cytochemical reactions

56. • Usefulness of techniques
• Identify functional subpopulations of
peripheral blood and lymphoid tissues
in health and disease.
• Classification of immunodeficiency
disorders
• Detection and classification of
lymphoid malignancies

57. • Characterization of leukemic
lymphocytes.
• Immunologic classification of
leukemia
• Showing phenotypic characteristics
of lymphoblasts and immature
lymphocytes

58. Quantitative Evaluation of
lymphocyte Populations
• T-lymphocytes predominate in the
thymus gland and peripheral blood,
thoracic duct lymph.
• B-lymphocytes predominate in bone
marrow and spleen.

59. • T-lymphocytes account for 70% of all
lymphocytes in peripheral blood
• B-lymphocytes constitute 20% of
peripheral blood lymphocytes.

60. • The remaining 10% are Null –
lymphocytes
• This distribution varies with species
and under various physiological
conditions and affected by techniques
of typing T or B-lymphocytes.
• The flow cytometer (fluorescent
activated cell sorter) is by far the
most accurate in computing lymphocyte
populations and sub-populations over
the rosetting methods.

61. • Flourescent antibody techniques using
antithymocyte antibody produce higher
results for T-lymphocytes that
rosetting methods.
• Peanut agglutinin (PNA) receptor is a
marker for cells in the thymic cortex
immature thymocytes – mice and man.
• It also marks bovine, ovine, caprine and
equine T-lymphocytes

62. • Surface Ig is a reliable marker for B
cells, but is labile and requires care
Ig marker is to be differentiated
from T-cells (few), macrophages,
suppressor macrophages, suppressor
cells (CD8+) and helper T-cells (CD8),
• CD8+ and CD4+ cells binds sIg via Fc
receptors.
• Rosetting B cells produces higher
values for B cells than B cells
identified by surface markers.

63. • T cell, and B cell populations in
peripheral blood and lymphoid tissues
vary with age and health.
• B cells are few in fetuses, steadily
increase postnatally to adult values.
• B cells, T cells and null cells decline
with age.
• Increased B cells and decreased T
cells indicate disease

64. • B cells increase in
• Bacterial diseases (eg mastitis).
• Leukemia
• Secretions from dry mammary glands
have increased T- lymphocytes.
• B cells are reduced in
• Acute lymphocytic leukemia (Also T
cells).

65. B Lymphocytes
• B lymphocytes constitute 20% of
circulating peripheral blood lymphocytes.
• B cells are identified by presence of
surface immunoglobulins and receptors for
complement C3b C3d.
• Complement receptors are absent in
plasma cells.
• Most human B lymphocytes have surface
IgM (monomeric) and IgD.
• Most canine lymphocytes have surface IgG
(70%).
• The rest have IgM.

66. • Human B cells exhibit B-cell specific
Ia like antigens on surface membrane.
Ia antigens are glycoproteins linked
genetically to major histocopatibility
complex with limited tissue
distribution.
• Ia-antigens occur also in
macrophages, early erythroid and
myeloid precursors.

67. • Human B cells rosette with mouse
erythrocytes.
• B-cells stain for 5’nucleotidase (only
in plasma membrane).
• B cells do not stain with acid
hydrolytic enzymes.
• Ia – like antigens are detected on
horse, cow, sheep, pig B-lymphocytes
by monoclonal antibodies.

68. T-Lymphocytes
• Peripheral blood lymphocytes have 21
– 85% T cells
• T cells are identified by E-rosetting
with heterologous erythrocytes
• T cells are also identified by
antithymocyte antibodies.
• Mature T cells form rosettes with
sheep erythrocytes (most species),

69. • T cells from canines form rosettes with
guinea pig and human type O erythrocytes
• Feline T-cells rosette with guinea pig, rat,
and mouse red cells.
• Porcine T cells rosette with rabbit
erythrocytes
• Equine T cells rosette with pig erythrocytes
• T-cells are also identified using specific
markers (surface markers).
• T-lymphocytes have receptors for Fc
portions of some immunoglobulins.
• Sub-sets of T lymphocytes with surface
receptors for IgG, IgM, and IgA are called
TG, Tm and TA cells.

70. • Tm cells reach up to 85% of all
lymphocytes, typically small to medium
sized, with smooth surface, poor
cytoplasmic organelles, no cytoplasmic
granules.
• These constitute helper T cells (CD4+)
• TG cells make 5-15% of T cells cells
• TG cells are large in size, contain
azurophilic granules, show villous surface,
abundant cytoplasm and well developed
organelles.
• TG cells constitute suppressor T cells
(commonly called CD8).

71. • T cells are also identified by mitogenic
responses to non specific stimulatory
agents such as plant lectins,
phytohemagluttinin (PHA) and
concanavalin (Con A).
• T cells are identified by cytochemical
markers acid hydrolytic enzymes, such
as acid phosphatase, β-glucuronidase,
acid α-naphthy acetate estarase – CD8
cells.

72. Pre-T, Pre-B and Null cells
• Pre-T and Pre-B cells have Tdt activity
• Pre-B cells show cytoplasmic IgM, without surface Ig
and complement receptors
• Mature B cells show cytoplasmic and surface Ig
including complement receptors.
• Null cells resemble morphologically and
cytochemically TG cells
• Null cell lack specific markers of T and B cells
• Null cells respond poorly to mitogens
• Null cells do not produce immunoglobulins in vitro
• Are non adherent and non phagocytic
• They are precursors of T and B cells
• Precursors of myeloid and erythroid cells

73. • Fig. 1: Human blood
• A two―lobed neutrophil (a) and a
small lymphocyte (b). There is
slight variation in size of the
normal erythrocytes. Wrights
stain x 400

74. • Fig. 3: Human blood: Nucleus of the neutrophil (a) has
not constricted into distinct lobes. Neutrophils with
more or less band‑form of nucleus are called band
neutrophils and are more immature than the lobed or
segmented neutrophils. Band neutrophils comprise 3 to
5 % of leukocytes in normal blood. Blood platelets (b)
and a small lymphocyte (c). Wrights stain x 400

76. • Human blood:
Blood smear
showing
erythrocytes,
platelets and
large
lymphocycte.
Wrights –
Giemsa x 400