Manual differential leukocyte count

Introduction:
A differential white cell count is the assigning of leucocytes to their individual categories, this
categorization being expressed as a percentage or, when the WBC is available, as an absolute
count..

Materials:
1- Clean, dust- free, grease-free glass slide (better to use glass slide with frosted end for
labeling).
2- Spreader (smooth end).
3- Fresh blood without anticoagulant or EDTA anticoagulated venous blood. (Do not use
heparinized venous blood because it leads to blue back ground).
4- Leishman’s stain

Methodology:
1- Make well-made well stained thin blood film.
2- Examine the smear systematically by using X10,X40 and X100.
3- Start counting in an ideal area (not more than 50% of RBCs are overlapped).
4- Classify at least 100 leukocytes either by battlement or longitudinal method.
5- Report results of the 100 cells classified as percentage.
6- Calculate the absolute leukocyte count by using the following formula:

Absolute leukocyte count= TWBCs count X relative count

100

1
 Reference values:

Absolute count Relative count

Neutrophils 2.0–7.0 × 109/l (40–75%)

Lymphocytes 1.0–3.0 × 109/l (20–45%)

Monocytes 0.2–1.0 × 109/l (2–10%)

Eosinophils 0.02–0.5 × 109/l (2–6%)

Basophils 0.02–0.1 × 109/l (<1–2%)

*WBC Correction for the Presence of Nucleated Red Blood Cells (nRBCs)