blood histology

1. Blood

2. • delivery of nutrients and oxygen directly or indirectly to cells
• transport of wastes and carbon dioxide away from cells
• delivery of hormones and other regulatory substances to and from cells
and tissues
• maintenance of homeostasis by acting as a buffer and participating
in coagulation and thermoregulation
• transport of humoral agents and cells of the immune system that
protect the body from pathogenic agents, foreign proteins, and
transformed cells (i.e., cancer cells)
Blood’s many functions include:

3. Blood cells and their derivatives include:
• erythrocytes, also called red blood cells (RBCs);
• leukocytes, also known as white blood cells (WBCs); and
• thrombocytes, also termed platelets.
The major plasma proteins include the following:
■ Albumin, the most abundant plasma protein, is made in the liver and serves
primarily to maintain the osmotic pressure of the blood.
■ α-Globulins and β-globulins, made by liver and other cells, include
transferrin and other transport factors; fibronectin; prothrombin and other
coagulation factors; lipoproteins and other proteins entering blood from tissues.
■ Ύ-Globulins, which are immunoglobulins (antibodies) secreted by plasma cells
in many locations.
■ Fibrinogen, the largest plasma protein (340 kD), also made in the liver, which,
during clotting, polymerizes as insoluble, cross-linked fibers of fibrin that block
blood loss from small vessels.
■ Complement proteins, a system of factors important in inflammation and
destruction of microorganisms.

4. The volume of packed erythrocytes in a sample of blood
is called the hematocrit (HCT) or packed cell
volume (PCV). The hematocrit is measured by centrifuging
a blood sample to which anticoagulants have been added,
and then calculating the percentage of the centrifuge tube
volume occupied by the erythrocytes compared with that
of the whole blood.
A normal hematocrit reading is about 39% to 50% in men
and 35% to 45% in women; thus, 39% to 50% and 35% to
45% of the blood volume for men and women,
respectively, consists of erythrocytes.

6. ■ The liquid portion of circulating blood is plasma, while
the cells and platelets comprise the formed elements;
upon clotting, some proteins are removed from plasma
and others are released from platelets, forming a new
liquid termed serum.
■ Important protein components of plasma include
albumin, diverse α-and β-globulins, proteins of the
complement system, and fibrinogen, all of which are
secreted within the liver, as well as the immunoglobulins.

7. Erythrocytes
(a) Colorized SEM micrograph of normal erythrocytes with each side concave. X1800.
(b) Diagram of an erythrocyte giving the cell’s dimensions. The biconcave shape gives
the cells a very high surface-to-volume ratio and places most hemoglobin within a
short distance from the cell surface, both qualities that provide maximally efficient
O2 transport. Erythrocytes are also quite flexible and can easily bend to pass
through small capillaries.
(c) In small vessels red blood cells also often stack up in aggregates called rouleaux.
The standard size of RBCs allows one to estimate that the vessel seen is 15 mm in
diameter. X250.

8. The erythrocyte is an anucleated
cell in a shape of a biconcave
disccontaining hemoglobin. The surface
area of an erythrocyte is about 140 μm2
and its mean corpuscular (cell) volume
ranges from 80 to 99 femtoliter (1 fL =
10-15 L).
The life span of erythrocytes is approximately 120 days.

9. The normal concentration of erythrocytes in
blood is approximately 3.9 to 5.5 million
per microliter (μL, or mm 3 ) in women and 4.1-6.0
million/μL in men.

10. Inherited alterations in hemoglobin molecules are responsible for several
pathologic conditions, an example of which is sickle cell disease. This
disorder is caused by a mutation of one nucleotide (a point mutation) in
the gene for the hemoglobin β chain. The consequences of this single
substitution are profound. When the altered hemoglobin (called HbS) is
deoxygenated in capillaries, it polymerizes and forms rigid aggregates that
cause a characteristic sickle shape. The sickled erythrocyte is less flexible
and more fragile and has a shortened life span that can lead to anemia. it
increases the blood viscosity and can damage the wall of blood vessels,
promoting blood coagulation.
Sickle cells can block capillaries,
restricting O2 delivery to tissues and
leading to varying degrees of ischemia
or anoxia and organ damage.

12. ■ Red blood cells or erythrocytes, which make up the
hematocrit portion (~45%) of a blood sample, are enucleated ,
biconcave discs 7.5 μm in diameter, filled with hemoglobin for
the uptake, transport, and release of O2 , and with a normal life
span of about 120 days .
■ White blood cells or leukocytes are broadly grouped as
granulocytes (neutrophils, eosinophils, basophils) or
agranulocytes (lymphocytes, monocytes).

13. Leukocytes

15. ■ All leukocytes become active outside the circulation,
specifically leaving the microvasculature in a process
involving cytokines, selective adhesion, changes in the
endothelium, and transendothelial migration or diapedesis.
■ All granulocytes have specialized lysosomes called
azurophilic granules and smaller specific granules with
proteins for various cell-specific functions.

16. Neutrophils
(a) In blood smears neutrophils can be
identified by their multilobulated nuclei,
with lobules held together by very thin
strands. The cells are dynamic and the
nuclear shape changes frequently.
(b) Neutrophils typically have diameters
ranging from 12 to 15 μm.
The cytoplasmic granules are relatively
sparse and have heterogeneous staining
properties, although generally pale and
not obscuring the nucleus.
(c) Micrograph showing a neutrophil from
a female in which the condensed X
chromosome appears as a drumstick
appendage to a nuclear lobe (arrow).

17. A TEM of a sectioned human
neutrophil reveals the two types
of cytoplasmic granules:
the small, pale, more variably
stained specific granules (S) and
the larger, electron-dense
azurophilic granules (A).
Azurophilic granules are modified
Lysosomes.
The nucleus (N) is lobulated and
the central Golgi apparatus (G)
is small. Rough ER and
mitochondria are not abundant,
because this cell utilizes glycolysis
and is in the terminal stage of its
differentiation. X25,000.

18. With electron microscopy, neutrophils have three distinguishing features. Firstly,
multiple nuclear lobes N with condensed chromatin; these lobes are seen as separate
in the thin EM sections. Secondly, the cytoplasm contains many membrane-bound
granules. The primary granules P are large, spheroidal and electron-dense. The
secondary or specific granules S are more numerous, small and often rod-like and are
of variable density and shape.
Neutrophils are highly motile cells. Motility and endocytotic (phagocytic) activity
are reflected in a large content of the contractile proteins, actin and myosin, as well as
tubulin and microtubule-associated proteins.

19. Neutrophil phagocytosis

20. As an incidental finding the inactivated X
chromosome in females is seen as
a small drumstick-shaped appendage D
in a few (3%) percent of neutrophils.
The lifespan of a neutrophil is a few
days and they are rarely found in
normal tissue.

21. Neutrophils are highly motile, phagocytose bacteria and kill them by fusing
the phagosome with neutrophil primary granules and producing activated
oxygen derivatives. Under certain conditions, they degranulate, releasing
granule contents including inflammatory mediators, antibacterial enzymes
and tissue matrix breakdown enzymes. Massed neutrophils and their debris
in tissue are visually recognised as pus.

22. ■ Neutrophils, the most abundant type of
leukocyte, have polymorphic, multilobed nuclei,
and faint pink cytoplasmic granules that contain
many factors for highly efficient phagolysosomal
killing and removal of bacteria.

23. Eosinophils
Eosinophils are about the same size
as neutrophils but have bilobed
nuclei and more abundant coarse
cytoplasmic granules.
The cytoplasm is often filled with
brightly eosinophilic specific
granules, but it also includes some
azurophilic granules.
(a) Micrograph shows an eosinophil (E) next
to a neutrophil (N) and a small lymphocyte
(L).
(b) Even with granules filling the cytoplasm,
the two nuclear lobes of eosinophils are
usually clear.

24. Ultrastructurally a sectioned
eosinophil clearly shows the
unique specific eosinophilic
granules (EG), as oval structures
with disc-shaped electron-dense,
crystalline cores.
These granules, along with a few
lysosomes and mitochondria (M),
fill the cytoplasm around the
bilobed nucleus (N). X20,000.

25. Eosinophils circulate in the blood for approximately 18 hours
and exit from capillaries to enter the tissues, where the majority
(>95%) of eosinophils reside.
Eosinophils are believed to survive in tissues for extended periods
(8-12 days and longer), but experimental data are limited.
Eosinophils do not generally recirculate; from the intestine they
exit into the bowel lumen or otherwise undergo lysis.

26. Most cells have a bilobed nucleus, but as cells mature in
tissue, the nucleus can further segment. The densely packed
cytoplasmic granules may partially obscure the nucleus.

27. Electron micrograph of a human eosinophil. The nucleus is bilobed,
but the connecting segment is not within the plane of section. The
granules are of moderate size, compared with those of the basophil,
and show a crystalline body (Cr) within the less electron-dense matrix
of the granule. M, mitochondria. X 26 000.

28. On electron microscopy, these cells
are dominated by the large, ovoid,
specific granules S, each containing
an elongated crystalloid. In humans,
the crystalloids are relatively
electron-lucent and irregular in
form, but in many other mammals
they have a more regular discoid
shape. Eosinophils have only small
numbers of mitochondria and
extensive smooth and some rough
endoplasmic reticulum SER and RER.
Note also the free ribosomes R and
the bilobed nucleus N.

29. ■ Eosinophils have bilobed nuclei and
eosinophilic specific granules containing factors
for destruction of helminthic parasites and for
modulating inflammation.
The cytoplasm of eosinophils contains two types
of granules: numerous, large, elongated specific
granules and azurophilic granules (otherwise, the
eosinophil contains only a sparse representation
of membranous organelles).

30. Basophils
Basophils are also approximately the same size as neutrophils and
eosinophils, but they have large, strongly basophilic specific granules that
usually obstruct the appearance of the nucleus which usually has two
large irregular lobes.

31. A TEM of a sectioned basophil reveals the single bilobed nucleus (N) and
the large, electron-dense specific basophilic granules (B). Basophils exert
many activities modulating the immune response and inflammation and
have many functional similarities with mast cells, which are normal, longer-
term residents of connective tissue. X25,000.

32. Basophils, the rarest type of circulating leukocyte,
have irregular bilobed nuclei and resemble mast cells
with strongly basophilic specific granules containing
factors important in allergies and chronic inflammatory
conditions, including histamine, heparin, chemokines,
and various hydrolases.

33. Lymphocytes
Lymphocytes are agranulocytes and lack the
specific granules characteristic of
granulocytes. Lymphocytes circulating in
blood generally range in size from 6 to 15 μm
in diameter and are sometimes classified
arbitrarily as small, medium, and large.
The most numerous small lymphocytes shown
here are slightly larger than the neighboring
erythrocytes and have only a thin rim of
cytoplasm surrounding the spherical nucleus.
X1500.

34. Medium lymphocytes are distinctly larger than erythrocytes.
Large lymphocytes, much larger than erythrocytes, may represent
activated cells that have returned to the circulation.

35. Ultrastructurally a medium-sized lymphocytes is seen to be mostly
filled with a euchromatic nucleus (N) surrounded by cytoplasm
containing mitochondria (M), free polysomes, and a few dark
lysosomes (azurophilic granules). X22,000.

36. The lymphocytes circulate between
various lymphoid tissues and other
tissues of the body via the blood and
lymphatic vessels. They continuously
transit through tissues and back into the
circulation as part of immune
surveillance.
Lymphoid cells have a variable lifespan
ranging from weeks to an indefinite
lifespan and, unlike granulocytes, are not
end cells; they can proliferate, with most
proliferation occurring in tissues.

37. Lymphocytes constitute 20% to 40%
of the circulating leukocytes, with 1.0
to 4.5 × 10 9 /L. Lymphocytes are
the smallest of the white cells, being
only slightly larger than erythrocytes. They
generally have a round or oval, densely
stained nucleus with clumped chromatin
and a relatively small amount of pale,
basophilic, non-granular cytoplasm.
These small lymphocytes are ‘inactive’
forms.

38. The punctate appearance of the cytoplasm is caused by the presence of numerous
free ribosomes. Several mitochondria (M) are evident. The cell center or centrosphere
region of the cell (the area of the nuclear indentation) also shows a small Golgi
apparatus (G) and a centriole (C ). X 26000.
Electron micrograph of a medium-sized lymphocyte.

39. ■ Lymphocytes, agranulocytes with many
functions as T- and B-cell subtypes in the immune
system, range widely in size, depending on their
activation state, and have roughly spherical
nuclei with little cytoplasm and few organelles.

40. Monocytes
Monocytes are large agranulocytes with
diameters from 12 to 15-20 μm that
circulate as precursors to macrophages
and other cells of the mononuclear
phagocyte system.
Micrographs of monocytes showing their
distinctive nuclei which are indented,
kidney-shaped, or C-shaped.

41. Ultrastructurally the cytoplasm
of a monocyte shows
a Golgi apparatus (G),
mitochondria (M), and
lysosomes or azurophilic
granules (A).
Rough ER is poorly
developed and there are some
free polysomes (R). X22,000.

42. Monocytes constitute from 2% to 10% of leukocytes in peripheral
blood. They circulate for 3 to 4 days on average before migrating into
tissues. These cells are motile, highly phagocytic and may mature in
tissues into tissue resident macrophages of varying kinds with
extended lifespans.

43. Monocytes are characterised by a large, eccentrically placed nucleus which
stains less intensely with more open chromatin than other leucocytes.
Nuclear shape is variable but often with a deep indentation in the nucleus
near to the centre of the cell, giving a horseshoe shape. Two or more
nucleoli may be visible. Cytoplasm is abundant and stains pale greyish-blue
with Romanowsky methods. There are numerous small, purple-stained
lysosomal granules and cytoplasmic vacuoles which confer a ‘frosted-glass’
appearance.

44. The cytoplasm contains a variable
number of ribosomes, polyribosomes
and little rough endoplasmic reticulum.
The Golgi apparatus G is well developed
and is located with the centrosome in
the vicinity of the nuclear indentation.
Small elongated mitochondria M are
prolific. Small pseudopodia P extend
from the cell, reflecting phagocytic
ability and amoeboid movement.
The cytoplasmic granules Gr of
monocytes are electron-dense and
homogeneous. Half resemble primary
(azurophilic) granules of neutrophils
and these contain myeloperoxidase,
acid phosphatase, elastase and cathepsin
G. The other half are secretory granules
containing plasma proteins, membrane
adhesion proteins and tumour necrosis
factor alpha (TNF-α).

45. Electron micrograph of a human mature monocyte.
The nucleus is markedly indented, and adjacent to this site,
a centriole (C ) and several Golgi profiles (G) are evident. The
small dark granules are azurophilic granules, the lysosomes (L)
of the cell. The slightly larger and less dense profiles are
mitochondria (M). X 22 000.

46. ■ Monocytes are larger agranulocytes with
distinctly indented or C-shaped nuclei that
circulate as precursors of macrophages and
other cells of the mononuclear phagocyte system.

47. Platelets
Platelets are cell fragments 2-4 μm in diameter derived
from megakaryocytes of bone marrow. Their primary
function is to rapidly release the content of their granules
upon contact with collagen (or other materials outside of
the endothelium) to begin the process of clot formation
and reduce blood loss from the vasculature.

48. In a blood smear, platelets (arrows) are
often found as aggregates. Individually
they show a lightly stained hyalomere
region surrounding a more darkly
stained central granulomere containing
membrane-enclosed granules.

49. Ultrastructurally a platelet shows a
system of microtubules and actin
filaments near the periphery, called
the marginal bundle (MB), which is
formed as the platelet pinches off
from megakaryocyte, and helps
maintain its shape. An open
canalicular system (OCS) of
invaginating membrane vesicles
continuous with the plasmalemma
facilitates rapid degranulation
upon activation and Ca 2+ release.
The central granulomere region
contains small dense delta granules
(δG), larger and more numerous
alpha granules (αG), and glycogen (G).
X40,000.

50. Platelets are small, round or oval, biconvex cytoplasmic discs, varying in size from
1.5-2 to 3.5-4 µm diameter. In blood smears, they have a central purple-stained
granular appearance, due to their numerous organelles and a poorly seen pale-
staining periphery. Platelet numbers in circulating blood range from 150 to 500 ×
Billion/L, and they survive there for 5 to 10 days.

51. Platelets have most of the organelles of
other cells except nuclei. The conspicuous
granules/organelles when seen on EM
can be classified into several types:
• Alpha granules are variable in size and
shape. These contain many proteins
related to adhesion, blood clotting and
growth factors for repair.
• Dense granules are very electron-dense.
They contain serotonin, ADP, ATP, Ca 2+
and Mg 2+ .
• Lysosomes are membrane-bound
vesicles as found elsewhere, containing the
usual enzymes.
Platelets are surprisingly complex. They
have a marginal band of microtubules in
their peripheral cytoplasm, associated
with abundant contractile proteins, actin
and myosin; this is a contractile system.

52. Platelets are functionally complex. They have over 50 different
types of surface receptors. They respond to vessel injury to
prevent bleeding and are active in blood clotting and tissue repair.
On exposure to damaged tissue, platelets adhere to exposed
collagen and other basement membrane proteins via their surface
membrane receptors. Activation leads to contraction of the
microtubule system and degranulation with release of granule
contents, serotonin and ADP. Activated platelets also produce the
lipid mediator thromboxane. These signals recruit adherence of
additional platelets, with formation of a platelet plug.

53. Platelet electron micrograph and diagram. a. High-magnification electron micrograph of a
platelet situated between an erythrocyte on the left and an endothelial cell on the right. Visible
organelles include a mitochondrion, microtubules, a single profile of the surfaceconnected open
canalicular system, profiles of the dense tubular system, the moderately dense granules, a
single very dense granule, and glycogen particles. The microfilaments are not evident against
the background matrix of the platelet. b. Diagram of a platelet showing the components of the
four structural zones.

54. Disorders of platelets
Reduced platelet numbers is called thrombocytopenia. Low
levels, especially <20 × 10 9 /L, are associated with spontaneous
small vessel bleeding (petechiae) usually in skin and in the
bowel wall; this is a life-threatening situation.
There are many genetic disorders with mutations in various
proteins affecting platelet function. The commonest is von
Willebrand disease, which is due to defects in von Willebrand
factor (FVIII-VWF), a complex adhesion molecule produced in
endothelium and megakaryocytes.
Drugs can affect platelet function. Aspirin (acetylsalicylic
acid) blocks the enzyme cyclooxygenase, inhibiting thromboxane
production, and thereby impairing platelet function.

55. Platelet aggregation, degranulation, and fibrin clot formation.
Minor trauma to vessels of the
microvasculature is a routine occurrence
in active individuals and quickly results in
a fibrin clot, shown here by SEM. Upon
contact with collagen in the vascular
basement membrane, platelets (P)
aggregate, swell, and release factors that
trigger formation of a fibrin meshwork (F)
that traps erythrocytes (E) and more
degranulating platelets. After repair of the
vessel wall, fibrin clots are removed by
proteolysis due primarily to locally
generated plasmin, a nonspecific
protease. X4100.

56. Scanning electron micrograph of blood
clot. High-magnification scanning
electron micrograph shows initial stage
of blood clot formation. Red blood cells
are entrapped in a loose mesh of fibrin
fibers that are extensively cross-linked to
form an impermeable hemostatic plug
that prevents movement of cells and
fluids from the lumen of the injured
vessel. X1 600.

57. ■ Platelets are small (2-4 μm) cell fragments
derived from megakaryocytes in bone marrow,
with a marginal bundle of actin filaments, alpha
granules and delta granules, and an open
canalicular system of membranous vesicles; rapid
degranulation on contact with collagen triggers
blood clotting.

58. ~7.5 µm diameter

59. 2 to 4 µm diameter

60. have distinctive nuclei with 2 to 5 lobes (polymorphic)
and azure granules. Specific granules are unstained.

61. contain distinctive large, eosinophilic granules.
Nuclei usually have two lobes.

62. rare cells with distinctive large, basophilic granules.

63. (6 to 9 µm diameter) with a thin rim of cytoplasm. Nuclei are
round and densely stained heterochromatin.

64. larger cells (9 to 15 µm diameter) with more
cytoplasm than small lymphocytes. Nuclei
contain a mixture of euchromatin and
heterochromatin.

65. large cells (12 to 20 µm diameter) with "kidney-shaped" or
notched nuclei with chromatin having a "raked" appearance. The
cytoplasm is bluish-gray.

66. The formed element indicated by the arrow:
1. comprises less than 1% of circulating leukocytes.
2. produces platelets.
3. is derived from a lymphoid progenitor cell.
4. can leave the bloodstream and differentiate into a tissue macrophage.
5. produces IgE antibodies