DIPLOMA LECTURE SERIES

THE CONSTITUENTS OF BLOOD AND THE

INTERPRETATION OF HAEMATOLOGICAL INDICES
Blood consists of plasma cells (red cells, white cells and platelets)
The blood is primarily a medium for the carriage of O2, nutrient

materials
Hormones
Anti- infective agents (e.g. antibodies) to the tissues
Removal of CO2
Other waste products
From the tissues and their elimination from the body. Blood has
almost ubiquitous distribution in the body with unique chemical
characteristics, making it a most efficient transport system.
Properties of haemoglobin (Hb) allow the carriage of large quantities
of O2 needed for metabolic activities.
Buffering power of Hb is also an important factor in maintaining
constancy of blood PH.
Plasma proteins exert an oncotic pressure that influences exchange
of fluid between the blood and the tissues. Proteins also combine with
many substances e.g. iron, thyroxin, and steroid hormones, to form
transportable complexes from which the active components are
released at the appropriate sites.
Plasma and platelet contain all the factors required for clotting. Thus,
loss of blood, as in injury, is reduced by inherent properties of the
blood itself.

THE CONSTITUENTS OF BLOOD AND THE

INTERPRETATION OF HAEMATOLOGICAL INDICES
(Contd)
Anti
bodies belong to the gamma globullins. They are essential for resistance to

infections.
Haemagglutinins
are genetically important. Also relate to blood transfusion.
Agglutinogens
Leucocytes
partake in inflammatory reactions
Plasma
The Plasma Proteins [ 6.4-8.3g/100ml ]
Albumin
two principal groups conventionally recognized.
Globulins
Globulin subdivides into fraction: 1, 2, and fibrinogen
Average normal concn are in g/100ml
Albumin 4.8
Globulins 2.3
Fibrinogen 0.3
Globulin electrophoresis isolates a number of proteins with specific physiological
functions
e.g.
Prothrombin
Plasma thrombo plastin
Angiotensinogen
Immune globulins
Anterior pituitary hormones.

INTERPRETATION OF HAEMATOLOGICAL INDICES

Many haematological (and other) diagnoses are made by careful/appropriate

examination of the peripheral blood film. It is also necessary for

interpretation of the FBC indices.
Anisocytosis: variation in RBC size e.g. megaloblastic anaemia; thalassaemia,

IDA [Fe deficiency anaemia]

Acanthocytes: Rbcs with many spicules
unstable rbc membrane structure as in abetalipo proteinemia.
Basophilic rbc Stippling: Denatured RNA in RBC accelerated

erythropoiesis/defective Hb synthesis e.g. Pb poisoning; megaloblastic

anaemia; myelodysplasia; liver dx; Hb opathy e.g. thalasaemias
Blasts: nucleated precursor cells.
Not normal feature in peripheral blood.
Seen in e.g. myelofibrosis; leukaemia; malignant infiltration by carcinoma.
Burr Cells:: irregularly shaped RBCs seen in uremia
Dimorphic Picture 2 populations of rbc. Seen after Px for fe, B12 or

folate
Mixed deficiencies (fe + B12 or folate) post transfusion
Sidero blastic anaemia

INTERPRETATION OF HAEMATOLOGICAL INDICES (Contd)

A clone of abnormal erythroblasts produce abnormal cells, alongside normal rbc production.
Howel-Jolly bodies DNA nuclear remnants in rbc which are normally removed by the spleen
E.g Post-Splenectomy
Hyposplenism (eg SCD, celiac dx, congenital uncreative colitis/crohns;
Myeloproliferative dx
Amyloid
Also in dyserythropoietic states: myelodysplasia; megaloblastic anaemia
HyPoChromia: less dense staining or rbcs due to Hb synthesis e.g. Fe-def-anaemia; sidero

blastic anaemia (Fe stores unstable).

Left Shift immature neutrophils from the marrow e.g. infection.
Leucoerythroblastic Anaemia: immature cells (myelocytes, promyelocytes, metamyelocytes,

normoblasts)in the film.

Eg marrow infiltration as in malignancy
Anorexia
Sepsis
Severe haemolysis
Leukaemoid Reaction marked leucocytosis
(wcc > 50x109/L)
Eg severe illness as in infection
Burns
Leukaemia

INTERPRETATION OF HAEMATOLOGICAL INDICES (Contd)

Pappenheimer Bodies Granules of siderocytes containing iron. Eg lead poisoning
Carcinomatosis, Post-splenectomy
Poikilocytosis = Variation in rbc shape . Eg IDA, Myelofibrosis, Thalasaemia
Polychromasia Rbcs of different ages stain unevenly (young are more bluish).
Eg bleeding
Haematinic replacement (Fe S04; B 12, folate)
Haemolysis
Marrow infiltration
Retic count.
RETICULOCYTES: (normal: 0.8-2% or < 85x105/L)
Young, larger rbcs (containing RNA) signifying active erythropoiesis
Eg haemolysis
Haemorrhage
If B12, Fe or folate is given to marrow that lacks these.
Right Shift: Hypermature white cells: hypersegmented polymorphs (>5 lobes of

nucleus)
Eg megaloblastic anaemia
Uraemia
Liver dx

INTERPRETATION OF HAEMATOLOGICAL INDICES (Contd)

Rouleaux Formation : rbcs stack on each other ESR
Eg chronic inflammation
Para protienemia
Myeloma.
Spherocytes: Spherical cells
Eg Hereditary spherocytosis; autoimmune haemolytic anaemia
Schistocytes: Fragmented rbcs slice by fibrin bands in

intravascular hemolysis
Eg microangio pathic anaemia as in DIC, haemolytic uraemic
syndrome, thrombotic thrombo-cyto penic purpura, preeclampsia
Target Cells (Mexican hat cells)
Rbcs with central staining, a ring of pallor and an outer rim of
staining
Eg liver dx
Hyposplenism
Thalalsaemia
IDA.

INTERPRETATION OF HAEMATOLOGICAL
INDICES (Contd)
The Differential White Cell Count: Neutrophil 2-7.5x109/ [40-75% ]

but absolute values are more meaningful than percentages.

Increase: Bacterial infections
Inflammation e.g. myocardial infarction
Polyarteritis nodosa
Myelo proliferative disorders
Drugs (steroids)
Disseminated malignancy
Stress of trauma, surgery, burns; haemorrage, seizure.
Decrease (Neutropenia)
Viral infections
Drugs e.g. post-chemopx,cytotoxics, sulfonamides; carbimazole
Severe sepsis
Neutrophil antibodies (SLE, haemolytic anaemia) destruction
Hypersplenism e.g. feltys syndrome
Bone marrow failure - production

INTERPRETATION OF HAEMATOLOGICAL INDICES

(Contd)

Lumphocytes 1.5-4.5x109/L (20-45%)

Increase: acute viral infections; chronic infections eg TB, Brucella hepatitis,

syphilis
Leukaemia and lymphomas eg CLL
Decrease:
steroid Px, SLE, Uraemia
Legion naires dx, HIV infection, Marrow infiltration
Post-chemo Px or radio Px. T-lymphocyte subset CD 4 count in HIV
EOSINOPHILS 0.04-4x109/L (1 6%)
Increase: Drug reactions eg with erythema multiforme
Allergies, asthma, atopy
Parasitic infections (esp invasive helminths).
Skin dx esp. pemphigus, eczema, psoriasis, dermatitis herpetitoformis
Malignant dx eg lymphomas and eosinophilic leukaemia.
Adrenal insufficiency, irradiation, lofflers syndrome, during the
convalescent phase of any infection.
The hyper eosinophilic syndrome is a disease of unknown cause with
sustained eosinophilic count > 15x109/L for more than 6wks, leading to
end-organ damage (endomyocardial fibrosis causing restrictive
cardiomyopathy, skin lesions, thromboembolic dx, pulmonary dx,
neuropathy and hepatosplenomegaly).

INTERPRETATION OF HAEMATOLOGICAL
INDICES (Contd)
MONOCYTES

0.2-0.8x109/L (2-10%)

Increase:

post-chemo px or radiotherapy, chronic

infections (e.g. malaria, TB, brucellosis, protozoa) malignant
dx, myelodysplasia.

Basophils 0-0.1x109/L (0-1%)

Increase:

myeloproliferative dx, viral infection IgE

mediated hypersensitivity reactions (eg urticaria,
hypothyroidism) inflamm disorders eg rheumatoid arthritis.

MACROCYTOSIS (MCV> 96fL)

Often due to alcohol excess without any accompanying

anaemia. Only 5% are due to B12 deficiency

B12 and folate deficiency are megaloblastic anaemias.

A megaloblast is a cell in which nuclear maturation is

delayed compared to the cytoplasm. This occurs with B12

and folate deficiency, as they are both required for DNA,
synthesis.

INTERPRETATION OF HAEMATOLOGICAL INDICES

(Contd)
Causes of Macrocytosis
Megaloblastic: B12 and folate citotoxic drugs.
Non-megaloblastic: ROH, reticulocytosis

liver dx

pregnancy

hypothyroidism
Others:
myelodysplasia
myeloma
myeloproliferative dx
aplastic anemia
Tests: B12 and folate result in similar blood film and
bone marrow biopsy appearances.
Blood film hypersegmented polymorphs in B12 and
folate ; (target cells if liver dx).

INTERPRETATION OF HAEMATOLOGICAL INDICES

(Contd)
Pernicious Anaemia

Caused by an autoimmune atrophic gastritis leading to achlor

hydria and lack of gastric intrinsic factor secretion.

Test:Hb (3-11gIdL)
MCV
WCC & platelets in severe cases
Serum B12
Retics or normal.
An approach to haemolytic anaemia
Haemolysis is the premature breakdown of rbcs before their
normal life span of about 120 days. Intravascular when it
occurs in circulation; etravascular when it occurs in the
reticulo endothelial system ie macrophages of liver, spleen
and bone marrow. In sickle cell anaemia lifespan may be as
short as 5 days. Haemolysis may be asymptomatic when the
bone marrow compensates sufficiently.
The approach is to first confirm hemolysis then find the cause.

INTERPRETATION OF HAEMATOLOGICAL INDICES

(Contd)
If increased rbc breakdown, there will be :
Anaemia with normal or MCV.
Bilrubin: unconjugated (prehepatic jaundice)
Urinary urobilinogen (no urinary conj, BR)
Serum lactic dehydrogenate (LDH) from rbc
If increased red cell production:
Reticulocytes causing MCV
Polychromasia.
If intravascular hemolysis: free plasma haemoglobin: from rbc
Methaem al buminaemia: some free Hb is broken down in

circulation haem + globin haem combines with albumin to make

met haemalbumin.
plasma haptoglobin: mops free Hb,and it is then removed by the

liver.

INTERPRETATION OF HAEMATOLOGICAL INDICES

(Contd)
Haemoglobinuria: causing red-brown urine, in absence of rbcs.
Haemosiderinuria: as haptoglobin binding capacity is exceeded

causing free Hb to be filtered by the renal glomerali, absorption of

free Hb via the renal tubules and storage in the tubular cells as
haemosiderin. This is detected in the urine in sloughed tubular cells
by Prussian blue stain 1wk after onset.
Thick + thin films may malaria.
Hypochromic microcytic anaemia (thalassaemia)
Sickle cells (sickle cell anaemia/SCD)
Schistocytes (microangiopathic haemogytic anaemia).
Abnormal cells haematogical malignancy
Spherocytes hereditary spherocytosis or autoimmune haemolytic
anaemia.
Elliptocytes hereditary elliptocytosis.
Heinz bodies G6 PD (bits cells)

INTERPRETATION OF HAEMATOLOGICAL INDICES

(Contd)
Further Tests
Direct antiglobulin (coombs) (DAT) Test indentifies rbc coated with
antibody or complement. If +ve immune cause of the haemolysis.
RBC lifespan determined by chromium labeling. Rarely done now.
Membrane abnormalities detected by osmotic fragility testing.
SICKLE CELL ANAEMIA
HbS polymerizes when deoxynated, causing RBCs to deform. This
produces sickle cells, which are fragile and haemolyse and also block
small vessels.
However, haemolysis is variable
Hb about 6-9 g/dL
retics 10-20%
BR
Film sickle cells
Target cells
Electrophoresis confirms the diagnosis and distinguishes SS,AS states
and other HB variants.
Target cells
Electrophoresis confirms the diagnosis and distinguishes SS,AS states
and other HB variants.

IN THE CASE OF BLEEDING DISORDER, WHAT

IS THE MECHANISM?
Prothrombin time (PT) tests the extrinsic system.
Expressed as ratio compared to control. INR (0.9-1.2)

abnormalities in factors I, II,V, VII, X

Prolonged by warfarin, VItK, liver dx, DIC.
Tested by adding thromboplastin to the blood.
Activated partial thromboplastin time (APTT) by adding

kaolin. Tests intrinsic system.

Abnormalities in factors I, II, V, VIII, IX, X, XI, XII. Normal =

35 45 secs.
Prolonged by heparin, haemophilia, DIC, liver

dx,haemophilia,factor V111 or 1X
Thrombin Time : [10 15 secs.] by adding thrombin to

plasma to convert fibrinogen to fibrin.

IN THE CASE OF BLEEDING DISORDER,

WHAT IS THE MECHANISM? CONTD
Prolonged by heparin, DIC, dysfibrinogenemia .
Bleeding Time Normal:[ < 7mins] is test of haemostasis by

making 2 small incisions into the skin of the forearm.

Raised in:ASA, platelet disorders,von Willibrand's dx
However, results are operator-dependent. So seldom used
now.
D-Dimers are fibrin degradation products, released from
cross-linked fibrin during fibrinolysis.
DIC, Venous thromboembolism as in DVT, pulm embolism
(PE).
May also be raised in infection and malignancy
Platelets: If low do: FBC, film, clotting time, PT. If prolonged
look out for liver dx or anticoagulant use.
If both PT and APTT are very raised with
platelets
D-dimers
Consider DIC
APTT:if prolonged, consider:liver dx,haemophilia[V111
or1x],heparin.

LEUKAEMIAS
Acute Myeloid Leukaemia (AML)
Blast cells derived from marrow myeloid elements. May be

few in the peripheral blood. So do marrow bx

WBC may be or normal.
Auer rods are diagnostic of AML but is now based on

immune phenotyping and molecular methods.

CML (chronic myeloid)
Philadephia chromosome (Ph)in >80%
Those with a ph have worse prognosis
WBC (>100X109/L) with whole spectrum of myeloid cells

ie neutrophils, myelocyte basophile, eosinophils, HB or

normal; platelets are variable.
Urate , B12
Neutrophil alk phos score
Bone marrow is hyper cellular
Ph found on cytogenic analysis of blood or bone marrow.

 CLL

(chronic lymphocytic)

lymphocytes
Hb
neutrophils
platelets
Marrow infiltration.
Later autoimmune haemolysis

HODGKINS LYMPHOMA
ESR or HB worse prognosis
LDH as it is released during cell turnover.

NON- HODGKINS LYMPHOMA

LDH worse prognosis

PANCYTOPENIA & MARROW FAILURE

Reduction in all the major cell lines: rbc, wbc and platelets.
marrow production as in aplastic anaemia, infiltration(eg

TB,acute leukaemia, myeloma, lymphoma, solid tumours),

megaloblostic anaemia.
Myelofibrosis, paroxysmal nocturnal haemoglobinuria, SLE.
Destruction as in hypersplenism.

MYLOPROLIFERATIVE DISORDERS
Polycyaemia rubra vera (PRV)
Rbc count, Hb, PCV
Often also Wbc, platelets
B12
Marrow shows hypercellularity with erythroid hyper plasia.
Neutrophil alk phos score ( in CML)
serum ery thropoietin
Red cell mass on

Cr studies with splenomegaly

Essential Thrombocythaemia
platelets often > 1000x109/L + abnormal functn
A clonal proliferation of megakaryocytes leading to
persistent elevation of platelets.
Myelomamalignant
A clonal proliferation of B-lymphocyte derived plasma cells.
monoclonal band or para protein on serum and/or urine
electrophoresis.
15

 FBC normocytic normo chronic anaemia.

film rouleaux formation
persistently ESR
Urea, creatinine, Ca2+
ALK phos normal.
Screening Test. Urine & urine electrophoresis, B-

microglobulin as prognostic test.

ERYTHROCYTE SEDIMENTATION RATE (ESR)

[20mm/hr]
Sensitive but non-specific indicator of the presence of disease.
Measures how fast rbcs fall through a column of
anticoagulated blood over 1 hour.
Certain proteins covering rbcs cause them to stick to each
other in columns, so, they fall faster (same phenomenon as
rouleaux).
ESR in any inflammation e.g.
Infection
Rheumatoid arth
Malignancy
Anaemia
Myocardial infarction
If slight elevation, repeat after 1month
Reassure patients with vague symptoms in whom there are no
pointers to specific disease.

But if very high > 100mm/h, there is 90% predictive value for
disease. Then consider FBC, plasma electrophosresis, U&E,
PSA chest & abd x-rays, bx bone marrow or temporal artery.
This is because in a survey, serious underlying dx later found
in such patients included myeloma, prostatic Ca, giant cell
arteritis, aortic aneurysm, leukaemia and lymphoma .
History + physical exam are very important. ESR also rises
with age. It is reliable to calcute:
for men upper limit of normal to be
Age (yrs)
2
Using Westergren method.
For women Age + 10
2
ESR may occur in-polycythemia (due to rbc concentration)
Sickle cell anaemia
Therefore, even a slight elevation in these patients prompts
further investigations.

THANKS FOR YOUR ATTENTION