Page 317 - The Veterinary Laboratory and Field Manual 3rd Edition

286 Susan C. Cork and Roy Halliwell

Total white cell count

PrEParE tHE dILutInG FLuId
Diluting fluid: 2% acetic acid in distilled
water coloured pale blue with Gentian vio-
let. 3

Prepare a 1/20 dilution of EDTA blood by adding
50 µl (0.05 ml) of blood to 950 µl (0.95 ml) of
diluting fluid in a mixing tube. Seal the tube and
mix by inversion and rotation. If a haemocytom-
eter pipette is used fill with EDTA blood to the
0.5 mark and then add diluent to the 101 mark.
Mix by rotating the pipette. Note that the pipette
has three markings on it, 0.5 mark in the middle
of the stem, 1 mark at the junction between stem
and bulb and 101 mark above the bulb. The total
volume of the pipette is 101 parts, of which one
Figure 5.3 Battlement counting method. Correctly part is in the stem and 100 parts in the bulb.
prepared blood smear with a think zone at one end Before loading the haemocytometer ensure
and a thin ‘feather’ at the other end (see also Figure that the diluted blood is well mixed and if using
5.2). the haemocytometer pipette, discard the first
few drops before loading the haemocytometer.
Leave the counting chamber for a few minutes
totaL cELL countS to allow the white blood cells to ‘settle’ before
counting. Count all the cells in the four large
For red and white cell counts a haemocytometer outer squares (see Figure 5.4b) (1 mm × 1 mm)
is required, preferably one with the Neubauer under the 10× or 20× objective. Include cells on
ruling (see Figures 5.4a and 5.4b). Use whole the left and upper lines but not cells on the lower
blood collected in EDTA anti-coagulant. Use and right lines. If nucleated (that is, immature)
calibrated pipettes with disposable tips or a red cells (nRBC) are encountered in mamma-
haemocytometer pipette for making the required lian blood smears, they should be recorded
dilutions. If a haemocytometer pipette is used separately from the leukocytes and reported as
it should be well washed and rinsed in distilled the number of nRBC/100 WBC. This will be
water after each use. Mouth pipetting is not rec- noted while performing the differential count as
ommended. As with most manual methods the they will not be able to be differentiated on the
following techniques are subject to a high degree haemocytometer.
of error especially when carried out by inexperi-
enced personnel.

Vet Lab.indb 286 26/03/2019 10:25