Experiment (3)
Suaad Mohammad(M.Sc)
The enumeration of the white blood cells
(The Total Leukocyte count)
A white blood cell count estimate the total number of white cells in a
cubic millimeter (mm3) of blood. It is important in the diagnosis of disease
especially when accompanied by a differential white cell count.
Apparatus and Reagents
1234-
Neubauers chamber with coverslip
A white cell pipette
Microscope
Diluting fluid
For WBC count, turk`s solution is used. This diluting fluid contain an acid
solution that lyses the RBCs and stain that stains the nuclei of the WBCs and
allows for easy identification and counting.
Turk`s solution
1- Glacial acetic acid
1 ml
2- Gention violet (1% aqueous solution) 2 ml
3- Distilled water
100 ml
Procedure
1- Clean the hemocytometer and its cover slip with an alcohol pad then
dry with a wipe.
2- Draw blood in a clean dry WBC pipette up to the mark 0.5 with all
possible accuracy.
3- Draw the diluting up to mark 11(dilution 1 in 10).
4- Mix the contents of pipette for minutes.
5- Dispel the first 4 drops of the contents.
6- Adjust the Neubauers chamber. It must be clean and dry. By holding
the cover slip between the fingers at the edges, place it in such
manner that both the ruled platforms are evenly covered by it. Load
it with the mixture, by holding the pipette at angle of 54 degree and
touching the space between the cover slip and the chamber by the
point of the pipette, an appropriate drop of the mixture is allowed to
run under the cover slip by capillary action.
7- Allow 2 minute for setting of cells then count.
Calculation
The total area of each the corner large squares is 1 mm2
Since the depth is 1/10 mm
So the volume of each corner large squares is 1mm2*1/10 mm = 1/10 mm3
The volume of the four corner large squares is
1/10 * 4 = 4/10 mm3
The number of WBCs in 1 mm3
X
The number of WBCs in 4/10
Y
Y*1
X=
1
=Y
4/10
= Y * 10/4 = Y * 2.5
4/10
Dilution is 20
X = Y * 2.5 * 20
So the number of WBCs in 1mm3 = number of cell counted * 50