LABORATORY INVESTIGATION OF

ANAEMIA

BY

PROF.YETUNDE. A. AKENOVA

Anaemia is an erythrocyte disorder characterized

by a haemoglobin concentration below normal for
the age and sex.
Anaemia can be categorized
1. A failure of production or shortened red cells
survival- Intrinsic or extrinsic defect.
2. Anaemia of chronic disease consequent on chronic
infection or inflammation or malignancy.

Haematocrit variation throughout life.

Term newborn (cord blood)
44 %
Term newborn (Capillary blood) 53 68
Infant (3 months) 30 38
Child (10 years)
37 44
Pregnancy (30 wks gestation)
26 34
Adult female
37 47
Adult male 42 54

Hct swings as high as 6-8% may occur with

correction of dehydration of volume overload.

How was the blood specimen obtain
Hgb/Hct values are frequently higher from a finger

stick than from a venous sample, unless

excessive pressure is applied to facilitate
blood flow.
Prolonged stasis from a tourniquet increases
the haematocrit as do muscular activity and
cold.

MCV = Hct RBC (Femtolitres)

MCH = Hgb RBC (Picograms)
MCHC = HgbHct ( g/dl RBC)
MCV is a measure of cell size
MCH is a measure of the average amount
of haemoglobin in each individual
cell.
MCHC Is a measure of the concentration of
haemoglobin in each cell.

Hgb (g/di)
Hct%
MCV (f1)
MCH (pg)
MCHC (g/dlRBCs)

Adult Male

Adult Female

14 18
42 54
82 98
27 32
31.5 36

12 16
37 47
82 98
27 32
31.5 36

PERIPHERAL SMEAR
COMMON CAUSES OF VARIOUS RBC ABNORMALITIES
Hypochromia

Iron deficiency

Microcytosis

Thalassaemia
Sideroblastic anaemia
Chronic inflammatory

Macrocytosis

Liver disease
Megaloblastic anaemia
Reticulocytosis
Myelodysplastic anaemia
Myelophthysis

Marked Amsocytosis
And Poikilocytosis

Marked iron deficiency

megaloblastic
Microanglopathic
Haemolysis
Leukoerythroblastosis
Haemoglobinopathies

Target cells

Liver disease
HaemoglobinC,AC, SS, SC
Post Splenectomy
Thalassaemia

Spiculated RBCs

Hereditary Acanthocytosis
Liver disease
Renal disease
Post Splenectomy
Hypothyroidism
Microangiopathic
Haemolysis

Tear drop cells

Leukoerythroblastosis
Megaloblastic anaemia
Thalassaemia
Erythroleukaemia

Howell Jolly bodies

Post Splenectomy
Megaloblastic Anaemia
Erythroleukemia

Spherocytes

Hereditary Sphenocytosis
Auto immune haemolysis
Haemoglobin C disorders CC,
SC.

Ovalocytes

Hereditary ovalocytosis
Megaloblastic anaemia
Iron deficiency
Thalassaemia.

1.Cell size Microcytic

Macrocytic
Normocytic
2. What is the basic Mechanism of the anaemia.
a. Decreased affective marrow production
b. Bleeding
c. Haemolysis
- Reticulocyte index used to assess the
bone marrow
response to the anaemia.
- The rate of Hct fall
Anaemia with appropriate reticulocyte response in the
absence of bleeding means haemolysis.

appropriateness of the

ANAEMIAS WITH LOW MCV

Iron deficiency
Thalassaemia
Anaemia of chronic disease
Some haemoglobinopathies
Sideroblastic anaemias
Aluminium toxicity.
INVESTIGATION
- History and Physical examination
- complete blood count serum iron
- serum iron
- Total iron binding capacity
- Serum Ferritin
- Serum transferin receptor

SPECIAL INVESTIGATION
GIT Radioraphic studies, Endoscopy, Gastroscopy,
Esophagoscopy,Exploratory laparotomy
Blood film
Anaemia with high MCV >100fl
Differential diagnosis
1. Liver disease
2. Alcoholism
3. No associated disease
4. Myelodysplastic syndromes
5. Drugs
6. Megaloblastic anaemias

 Serum Vit B12

RBC folic acid Assay
Bone marrow Aspiration

CAUSES OF VIT B12 DEFICIENCY IRON STAIN

Dietary, Gastric, Terminal Ileum disease, fish tape worm.
Pancreatic insufficiency
Drugs
Congenital (transcobalamin 11 deficiency)
HAEMOLYSIS AND BLEEDING
Anaemia with a Normal or slightly elevated MCV and an
appropriate Reticulocyte index.
Haemolysis would lead to Reticulocytosis Appropriate

bone marrow response

An anaemia with an appropriate reticulocyte

response in the absence of alert bleeding suggests
haemolysis.
Retic count in a Normal Hct 1%.
If haemolysis is chronic (months to years) the
marrow production may reach a level of production
greater than 10 times normal.
Reticulocyte = Reticulocyte X Pt hct
Normal hct

ALLOANTIBODY VERSUS AUTOANTIBODY

ALLOANTIBODY

Direct coombs Frequently negative.

AUTOANTIBODY

Positive

May be positive if
sensitized foreign
red cells are still
circulating.
Indirect Coombs Positive
Antibody Screen
(Panel)
specificity is seen

Positive or Negative

Panagglutination no
specificity.

ANAEMIA WITH A NORMAL MCV AND LOW

RETICULOCYTE INDEX
DIFFERENTIAL DIAGNOSIS
Renal failure
Anaemia of inflammatory disease (Anaemia of Malignancy.etc)
Mild (early) iron deficiency
Combined iron deficiency and megaloblastic anaemia
Sideroblastic anaemia
Aplastic anaemia
Bone marrow infiltration (myelophthisis)
Bleeding or hemolysis plus one of the above

Intravascular Haemolysis
When RBC destruction is rapid and occurs primarily Within
the vascular space, diagnosis is relatively easy.one sees the
following.
1. Haemoglobinaemia
2. Haemoglobinuria
3. Haptoglobin saturation
4. Haemosiderinuria

Extravascular haemolysis
1.
2.
3.
4.
5.
6.

Autoimmune haemolysis
Delayed haemolytic transfusion reactions
Haemoglobinopathies
Hereditary spherocytosis
Hypersplenism
Haemolysis with liver disease

The following cell types are seen

1. Spherocytes
2. Schistocytes
3. Spiculated cells
4. Bite cells

FLOW CHART FOR INVESTIGATING ANAEMIA

Low Haematocrit
(Haemoglobin)

*Corrected Reticulocyte Count

(Reticulocyte Index)
Normal value 0.5 2.5%
INVESTIGATE FOR:Folic Acid Deficiency
Vitamin B12 Deficiency
Liver Disease
Bone Marrow Failure
Sideroblastic Anaemia
Hypothyroidism
Chronic Alcoholism
Drugs

INVESTIGATE FOR:Haemolytic Anaemias

(See Chart II)

Elevated

Low / Normal
INVESTIGATE FOR:Macrocytes

Peripheral Blood Film

Microcytes

INVESTIGATE FOR:Iron Deficiency

Thalassaemia
Sideroblastic Anaemia
Anaemia of Chronic Disease

Reticulocyte Index = (Reti culocyte Count (%) x Pt. Haemoglobin (g/dl)

Expected Normal Haemoglobin (g/dl)
+ Correction Factor for Premature Release

Normocytes

Acute Blood Loss

Early Iron Deficiency
Infection
Inflammation
Neoplasia
Renal Disease
(Iron Folic Acid/B12)
Sideroblastic Anaemia
Red cell Aplasia
Aplastic Anaemia

+ Haemoglobin (g/dl)
10 11
79
<7

Correction Factor
1.5
2.0
2.5

ASSESSING HAEMOLYTIC ANAEMIAS

EXTRACORPUSCULAR
DISORDERS

INTRACORPUSCULAR
DISORDERS
SHOWS
Anisocytosis, Poikilocytosis,
Polychromasia, Nucleated Red
Blood Cells (NRBCs),
Fragmented Cells

Peripheral Blood Film

Osmotic Fragility

Acid Lysis (Hams Test)

Increased

Positive

Heredity Spherocytosis

Peripheral Blood Film

Direct Antiglobulin Test

Paroxysmal Nocturnal
Haemoglobin

Clotting Studies
Haemoglobin Electrophoresis

Abnormal

Positive

Haemoglobinopathies (e.g.
Hb S, Hb C)

Normal

Prolonged

Autoimmune
Haemolytic Anaemia

Disseminated Intravascular
Coagulation; Haemolytic Ureamic
Syndrome; Thrombotic
Thrombocytopenic Purpura

Normal

Red Cell Enzyme Assay

Family Studies

Deficient

Enzymopathies (e.g.
G6PD and PK Deficiency)

infection

Viral (e.g. CMV)

Bacterial (e.g. Clostridium welchi)
Parasite (e.g. Malaria)

Anaemia of chronic disease is characterized by

1. Low serum iron concentration and defective
incorporation into haemoglobin despite adequate
bone marrow stores of iron
2. A blunted erythropoietin response to anaemia.
When mild it is normocytic normochromic but it
can become severe and become hypochromic
microcytic
3. Absoulte reticulocyte count is reduced, also
ass.features indicative of chronic inflammation
may be present.e.g. Neutrophilia, thrombocytosis,
increased rouleaux formation.

Serum iron, serum transferin, are reduced while total

iron binding capacity is increased.
Serum ferritin is increased, increase ESR,
plasma viscosity, reduced albumin and increased
fibrinogen 2 macroglobulin and - globulins are
increased