Differential count of Leucocytes
Aim
To
perform differential counting of blood leucocytes
Principle
Blood
contains various types of cells such as erythrocytes, leucocytes and
platelets. The Leucocytes or WBCs exist
in two forms, granulocytes and agranulocytes. Granulocytes are further
classified as eosinophil, basophil and neutrophil, while agranulocytes are
lymphocytes and monocytes. These cells can be observed and identified based on
their cellular morphology and cytoplasmic staining characters in stained blood
smears.
The
neutrophil has a multilobed nucleus and a granulated cytoplasm. The granules are smaller and stains with both
acid and basic dyes; it is often called a polymorphonuclear leukocyte (PMN) for
its multilobed nucleus. The nuclei are
lobulated with thin bridges between the lobules.
The
eosinophil has a bilobed nucleus and a granulated cytoplasm that stains with
the acid dye eosin red. The cytoplasm stains faint pink and the granules are
large in red or red-orange colour.
The
basophils are few in number. They have a
lobed nucleus and heavily granulated cytoplasm.
The granules are large and dark or blue black in colour and fills the
cell.
Lymphocytes
have clear cytoplasm and dark violet coloured nucleus which almost fill the
entire cell.
Monocytes
are larger than lymphocytes and have a horse shoe shaped nucleus.
Procedure for Preparation of blood
smear and staining
A
clean glass slide was taken.
The
middle finger was scrubbed with alcohol and pricked using a sterile lancet.
A
drop of blood was placed on the slide at one end and the blood drop was spread
using another slide
The
smear was covered with Leishmann’s stain for 4 minutes. Same volume of ditilled water was added to
the stain in the slide and kept for another 10 minutes
The
slide was washed gently in running water, air dried and observed under
microscope using oil immersion objective.
Procedure for Differential counting
of leucocytes
The
slide was examined and different types of leucocytes were counted. 100 cells were counted following a serpentine
counting pattern, moving downwards and in chain like manner and various types
of leucocytes and their corresponding number per 100 leucocytes was tabulated.
This gives the differential count pattern of the blood smear.
Results
100
leucocytes were counted and the percentage of different granulocytes and agranulocytes
were determined. Their percentage
distribution was compared with the normal count. Higher or lower count of certain types of
leucocytes corresponds to pathological conditions. High neutrophil count indicate bacterial
infection, high eosinophil count occurs in allergic conditions or in parasitic
worm infections, high monocyte count indicates chronic infection, high
lymphocyte count indicates atigen- antibody interaction, etc.
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