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Unit 1 of 3 · B.Sc MLS Sem 6

Unit 1: Anaemias, leukaemias and cytochemistry

Applied Haematology-II notes · PTU syllabus (BMLS601-18)

3 min read8 topics10 exam questions
On this page
  1. Unit summary
  2. Classification of anaemia
  3. Iron deficiency anaemia
  4. Megaloblastic anaemia
  5. Haemolytic anaemias
  6. Classification of leukaemias
  7. Leukaemoid reaction
  8. Cytochemical staining
  9. Chromosomal studies in haematology
  10. Key terms
  11. Quick revision
  12. Important questions

Unit summary

Anaemias and leukaemias are the commonest serious blood disorders. This unit covers the classification and laboratory diagnosis of iron deficiency, megaloblastic and haemolytic anaemias, the classification and diagnosis of leukaemias, leukaemoid reactions, cytochemical stains and chromosomal studies.

After this unit you can

  • Classify anaemias and diagnose iron deficiency and megaloblastic anaemia
  • Diagnose haemolytic anaemias
  • Classify and diagnose leukaemias and distinguish leukaemoid reactions
  • Use cytochemistry and chromosomal studies

PTU syllabus topics

  • Classification and laboratory diagnosis of iron deficiency
  • megaloblastic and haemolytic anaemia
  • classification and laboratory diagnosis of leukaemias
  • leukaemoid reactions
  • cytochemical staining and its significance in haemopoietic disorders
  • chromosomal studies in haematological disorders
ComparisonClassifying anaemia by red cell size
MCV
Common causes

Microcytic hypochromic

Under 80 fL

Iron deficiency, thalassaemia

Normocytic

80–100 fL

Acute blood loss, chronic disease, haemolysis

Macrocytic (megaloblastic)

Over 100 fL

Vitamin B12 or folate deficiency

1

Topic 1

Classification of anaemia

  • Anaemia: haemoglobin below the reference for age and sex (WHO: below 13 g/dL in men, 12 g/dL in non-pregnant women, 11 g/dL in pregnancy).
ComparisonClassification
Morphological (by MCV)
Aetiological

Microcytic hypochromic

Iron deficiency, thalassaemia, anaemia of chronic disease, sideroblastic

Decreased production — deficiencies, marrow failure

Normocytic normochromic

Acute blood loss, chronic disease, kidney disease, aplastic anaemia, early haemolysis

Increased destruction — haemolysis

Macrocytic

Megaloblastic (B12, folate), liver disease, alcohol, hypothyroidism

Blood loss

2

Topic 2

Iron deficiency anaemia

  • Commonest anaemia in India — poor diet, hookworm, menstruation, pregnancy, chronic blood loss.
Key termsLaboratory findings
Blood counts
Low Hb, MCV, MCH, MCHC; high RDW
Smear
Microcytic hypochromic cells, pencil cells
Iron studies
Low serum ferritin (best test), low serum iron, high TIBC, low transferrin saturation (below 16%)
Marrow
Absent stainable iron (Perl's)
Response
Reticulocyte rise in 5–10 days on iron therapy
3

Topic 3

Megaloblastic anaemia

  • Causes: vitamin B12 deficiency (pernicious anaemia, vegetarian diet, ileal disease) and folate deficiency (diet, pregnancy, drugs such as methotrexate).
Key termsLaboratory findings
Blood counts
High MCV (often above 110 fL); pancytopenia
Smear
Oval macrocytes, hypersegmented neutrophils (shift to right)
Marrow
Hypercellular with megaloblasts (open chromatin, nuclear–cytoplasmic asynchrony); giant metamyelocytes
Biochemistry
Low serum B12 or folate; raised LDH and bilirubin (ineffective erythropoiesis); raised methylmalonic acid in B12 deficiency
Pernicious anaemia
Anti-intrinsic factor and anti-parietal cell antibodies
4

Topic 4

Haemolytic anaemias

ComparisonHaemolytic anaemias
Examples
Key tests

Membrane defects

Hereditary spherocytosis

Spherocytes; increased osmotic fragility; EMA binding test

Enzyme defects

G6PD deficiency

Bite cells, Heinz bodies; G6PD screening (fluorescent spot)

Haemoglobinopathies

Sickle cell, thalassaemia

Sickling test, HPLC

Immune

Autoimmune, haemolytic disease of newborn, transfusion reactions

Direct Coombs test positive

Non-immune acquired

Malaria, microangiopathy, PNH

Smear for parasites and schistocytes; flow cytometry for PNH

Key termsGeneral evidence of haemolysis
Increased destruction
Raised unconjugated bilirubin and LDH; low haptoglobin; urinary urobilinogen; haemoglobinuria in intravascular haemolysis
Increased production
Reticulocytosis, polychromasia, erythroid hyperplasia
5

Topic 5

Classification of leukaemias

ComparisonLeukaemias
Features
Laboratory

Acute myeloid leukaemia (AML)

Adults; blasts 20% or more

Myeloblasts with Auer rods; MPO positive

Acute lymphoblastic leukaemia (ALL)

Commonest childhood cancer

Lymphoblasts; MPO negative; PAS block positive; immunophenotyping (CD19, CD10)

Chronic myeloid leukaemia (CML)

Middle age; huge spleen

Very high WBC with all stages of myeloid cells; basophilia; low LAP score; Philadelphia chromosome t(9;22) BCR-ABL

Chronic lymphocytic leukaemia (CLL)

Elderly

Lymphocytosis with smudge cells; CD5 and CD23 positive

  • Classification systems: FAB (morphology, M0–M7, L1–L3) and WHO (morphology, immunophenotype, genetics); diagnosis confirmed by flow cytometry and cytogenetics.
6

Topic 6

Leukaemoid reaction

ComparisonLeukaemoid reaction vs CML
Leukaemoid reaction
CML

Cause

Severe infection, TB, burns, tumours

Clonal stem cell disease

WBC

Raised, usually below 50,000/µL

Often above 1,00,000/µL

Smear

Toxic granulation, Döhle bodies; few immature cells

Myelocyte and neutrophil peaks; basophilia

LAP score

High

Low

Philadelphia chromosome

Absent

Present

7

Topic 7

Cytochemical staining

ComparisonCytochemical stains
Stains
Significance

Myeloperoxidase (MPO)

Granulocytes and monocytes

Positive in AML; negative in ALL

Sudan black B (SBB)

Lipids in granules

Like MPO — myeloid

Periodic acid–Schiff (PAS)

Glycogen

Block positivity in ALL; erythroleukaemia

Non-specific esterase (NSE)

Monocytes, inhibited by fluoride

Monocytic leukaemias (M4, M5)

Leucocyte alkaline phosphatase (LAP)

Neutrophil enzyme

Low in CML; high in leukaemoid reaction and polycythaemia

Perl's Prussian blue

Iron

Iron stores, ring sideroblasts

8

Topic 8

Chromosomal studies in haematology

  • Conventional karyotyping of marrow cells (metaphases arrested with colchicine, G-banding), FISH and PCR detect translocations that diagnose and guide treatment.
Key termsKey abnormalities
t(9;22) BCR-ABL
CML; some ALL
t(15;17) PML-RARA
Acute promyelocytic leukaemia
t(8;21)
AML with good prognosis
Trisomy 21
Down syndrome — raised leukaemia risk
JAK2 V617F
Polycythaemia vera and other myeloproliferative neoplasms

Key terms

Ferritin
Iron storage protein; best marker of iron stores
Megaloblast
Large red cell precursor with delayed nuclear maturation
Haemolysis
Premature destruction of red cells
Auer rods
Needle-like inclusions in myeloblasts
Philadelphia chromosome
t(9;22) producing BCR-ABL

Quick revision

  • WHO anaemia cut-offs; morphological and aetiological classification.
  • Iron deficiency: low ferritin, high TIBC.
  • Megaloblastic: high MCV, hypersegmented neutrophils, low B12 or folate.
  • Haemolysis: membrane, enzyme, Hb, immune, acquired causes; general tests.
  • AML, ALL, CML, CLL; leukaemoid reaction; MPO, SBB, PAS, NSE, LAP, Perl's; key translocations.

Important exam questions

Practice questions written to the PTU exam pattern for this unit's syllabus: short answers (Section A style) and long answers (Sections B and C style).

Short-answer questions

  1. Q1.What is the best test for iron deficiency?
  2. Q2.Name two causes of megaloblastic anaemia.
  3. Q3.Which test shows immune haemolysis?
  4. Q4.What are Auer rods?
  5. Q5.How does LAP score help distinguish CML from a leukaemoid reaction?
  6. Q6.Which translocation defines CML?

Long-answer questions

  1. Q1.Describe the laboratory diagnosis of iron deficiency anaemia.
  2. Q2.Describe the laboratory diagnosis of megaloblastic anaemia.
  3. Q3.Describe the laboratory diagnosis of haemolytic anaemia.
  4. Q4.Classify leukaemias and describe their laboratory diagnosis.

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