LABORATORY EXAMINATIONS IN HAEMATOLOGY

RED BLOOD CELLS, WHITE BLOOD CELLS, AND PLATELETS

The key components of the blood examination consist of:
  - Red Blood Cells (RBC)
  - White Blood Cells (WBC)
  - Platelets (thrombocytes)

WHITE BLOOD CELL TYPES AND REFERENCE RANGE

The main types of white blood cells counted are:
  - Neutrophil: 55-70%
  - Lymphocyte: 20-40%
  - Monocyte: 2-8%
  - Eosinophil: 1-4%
  - Basophil: 0-2%

CONTENTS OF FULL BLOOD COUNT (FBC)

Full Blood Count (FBC)
Hemoglobin Concentration
Red Cell Count (RCC)
Mean Cell Volume (MCV)
Mean Cell Hemoglobin (MCH)
Mean Cell Hemoglobin Concentration (MCHC)
Hematocrit (Hct) or Packed Cell Volume (PCV)
Red Cell Distribution Width (RDW)
White Blood Cell - Total Count
WBC Differential Count
Platelet Count

FULL BLOOD COUNT (FBC)

Also known as Complete Blood Count (CBC).
Historical methods of blood counting included:
  - Manual hemoglobin concentration using colorimetric techniques.
  - Manual counts of red blood cells and white blood cells.
  - Manual platelet count.
Those parameters like MCV had to be calculated mathematically from other measured values (Hb, RCC, and PCV).

MODERN ANALYZERS

Modern blood analyzers utilize various methods such as:
  - Electronic impedance
  - Laser light scatter
  - Light absorbance
  - Staining characteristics
They provide a comprehensive range of CBC variables including:
  - Measured variables: Hb, PCV, RCC
  - Derived values: MCV, MCH, MCHC
  - Automated platelet counts and differential WBC counts.

BLOOD SAMPLE COLLECTION

Blood is typically collected from peripheral circulation.
Samples are placed in tubes containing EDTA (Ethylene Diamine Tetra-acetic Acid) acting as an anticoagulant to prevent clotting by chelating calcium.
Samples should be analyzed within 4 hours to ensure integrity.

PHLEBOTOMY TUBES

Types of tubes and their uses:
  - Purple or Lavender Stopper: For whole blood in CBC and blood smears; contains EDTA.
  - Light Blue Stopper: Contains sodium citrate, used for coagulation tests.
  - Gold or Black Stopper: Contains clotting additive; used for chemistry tests.
  - Green Stopper: Contains heparin, suitable for plasma determinations.
  - Grey Stopper: Contains glycolytic enzyme inhibitors for glucose determinations.
  - Red Top Tube: No additives; for antibody tests or drug testing.

MAIN BLOOD PARAMETERS MEASURED

Hemoglobin Concentration (Hb)
Red Cell Count (RCC)
Mean Cell Volume (MCV)
Mean Cell Hemoglobin (MCH)
Mean Cell Hemoglobin Concentration (MCHC)
Hematocrit (Hct) or PCV
Red Cell Distribution Width (RDW)
White Cell Count
WBC Differential Count
Platelet Count

HEMOGLOBIN (Hb)

Hemoglobin is found in red cells and is essential for oxygen transport from lungs to tissues.
Measurement Units: g/dL or g/L (SI units preferred in Europe; USA often uses g/dL).
Normal levels vary with age, gender, and altitude:
  - Children at Birth: 135-195 g/L
  - Children Age 2-5: 110-140 g/L
  - Children Age 6-12: 115-155 g/L
  - Adult Male: 130-180 g/L (13-18 g/dL)
  - Adult Female: 115-165 g/L (11.5-16.5 g/dL)
  - Pregnant Women: 110-138 g/L

GRADING ANAEMIA

Depending on Hb levels:
  - Not Anaemic: Hb > 12 g/dL (>120 g/L)
  - Mild Anaemia: Hb 10-11 g/dL (100-110 g/L)
  - Moderate Anaemia: Hb 8-9 g/dL (80-90 g/L)
  - Marked Anaemia: Hb 6-7 g/dL (60-70 g/L)
  - Severe Anaemia: Hb 4-5 g/dL (40-50 g/L)
  - Critical Anaemia: Hb < 4 g/dL (<40 g/L)

RED CELL COUNT (RCC)

Unit: × 10^12/L.
Normal Range:
- Male: 4.5-6.5 × 10^12/L
- Female: 3.8 - 5.8 × 10^12/L

CAUSES OF LOW RED CELL COUNT

Hypoproliferative anaemias (Iron, Vitamin B12, Folate deficiencies)
Aplasias (Idiopathic or drug-induced BM suppression)
Parvovirus B19 infection causing red cell aplasia.

CAUSES OF HIGH RED CELL COUNT

Polycythaemia
Thalassaemias (Increased production of microcytic, hypochromic cells)

HEMATOCRIT (Hct) OR PCV

Percentage of blood volume that is red blood cells.
Comparable to Packed Cell Volume (PCV).
To measure, blood is centrifuged in a microcapillary tube:
- Plasma occupies approximately 55% of whole blood.
- Erythrocytes occupy about 40-45% of blood volume.

TESTING FOR HCT

Measure length of total blood column in mm and the length of the red cell column:
- $PCV = rac{ ext{Length of red cell column (mm)}}{ ext{Length of total column (mm)}}$
Normal PCV reference ranges:
  - Children at Birth: 0.44–0.54
  - Children Age 2-5: 0.34–0.40
  - Children Age 6-12: 0.35–0.45
  - Adult Men: 0.40-0.54
  - Adult Women: 0.36-0.46

ANEMIA AND PCV

Decreased PCV indicates anaemia. Increased PCV may be found in:
- Plasma loss conditions (e.g., severe burns, dehydration).
- True polycythaemias.

RED CELL INDICES

Important indices for investigating anaemia include:
  - Mean Cell Volume (MCV)
  - Mean Cell Hemoglobin (MCH)
  - Mean Cell Hemoglobin Concentration (MCHC)
  - Red Cell Distribution Width (RDW)

MEAN CELL VOLUME (MCV)

MCV indicates the average size of red blood cells, measured in femtolitres (fL).
$MCV = rac{PCV (L/L)}{RBC (× 10^{12} / L)}$
Reference range: 80-98 fL
MCV Interpretation:
  - Decreased: Often in Iron deficiency, β-thalassaemia.
  - Normal: Blood loss or myelodysplasia.
  - Increased: Seen in B12/Folate deficiency, alcoholism, etc.

MEAN CELL HEMOGLOBIN (MCH)

MCH measures the average amount of hemoglobin in red blood cells (pg).
$MCH = rac{Hb (g/L)}{RBC (× 10^{12}/L)}$
Reference range: 27-32 pg

MEAN CELL HEMOGLOBIN CONCENTRATION (MCHC)

MCHC gives hemoglobin concentration in red blood cells.
$MCHC = rac{Hb (g/L)}{PCV (L/L)}$
Reference range: 310-370 g/L (31-37 g/dL)
Interpretation:
  - Low MCHC: Hypochromic in microcytic anaemias.
  - Normal MCHC: Macrocytic anaemias.
  - Elevated MCHC: Hyperchromic conditions.

RED CELL DISTRIBUTION WIDTH (RDW)

RDW reflects the variation in red cell size, indicating anisocytosis in a blood sample.
Determined using an automated counter, with normal values ranging from 11.5% - 14.5%.
RDW > 14.5% indicates increased variation (anisocytosis).
RDW < 11.5% indicates more uniform RBC sizes.

RETICULOCYTES

Immature red blood cells (RBCs) that indicate bone marrow activity.
Normal reticulocyte range: 0.5% - 2.5%.
Count increases in conditions such as:
  - Hemorrhage
  - Hemolysis
  - Response to therapy (iron, folate).
Count decreases in conditions like:
- Aplastic anemia
- Bone marrow infiltration.

DETECTION OF RETICULOCYTES

Stained with supravital dye (e.g., Methylene blue)
Modern blood counters can directly assess reticulocyte counts.

WHITE CELL COUNT (WBC)

Total WBC count is included in FBC, providing the number of WBCs in blood after lysing RBCs.
Whole blood is diluted in an acid reagent, preserving white cells.
Microscopic counts done via a haemocytometer, converting counts into standardized units.

WBC REFERENCE RANGE

Children (1y): 6.0-18.0 × 10^9/L
Children (4-7y): 5.0-15.0 × 10^9/L
Adults: 4.0-11.0 × 10^9/L

LEUKOCYTOSIS (HIGH WBC COUNT)

Causes include:
- Acute infections, inflammation, metabolic disorders, stress, traumas, and leukemias.

LEUCOPENIA (LOW WBC COUNT)

Causes include:
- Infections (viral, bacterial, parasitic), drug reactions, aplastic anaemia, and bone marrow infiltration.

WBC DIFFERENTIAL COUNT AND INTERPRETATION

Neutrophils (PMNs): 40-75% normal range, increased in infections and stress.
- Neutropenia: Mild decrease, and agranulocytosis for severe decrease.
Lymphocytes: 20-40% normal range, increased in viral infections, decreased during stress.
Eosinophils: 1-4% normal range, increased due to allergies, parasites, and certain diseases.
Basophils: 0-2% normal range, increased in conditions like leukemia.
Monocytes: 2-8% normal range, increased in infection and recovery phases.

PLATELET COUNT

Platelets are crucial for blood clotting, with a reference range of 150-350 x 10^9/L.
Factors affecting platelet levels include:
- Thrombocytopenia: Reduced production or increased destruction.
- Thrombocytosis: Can be primary (clonal) or reactive (secondary) due to various conditions.