Page 319 - The Veterinary Laboratory and Field Manual 3rd Edition

288 Susan C. Cork and Roy Halliwell

Physiological salt solution (0.85%): 8.5 g 5.4 red cell indices and anaemia
sodium chloride (NaCl) in 1 l of distilled
water. Red cell indices include the measurement of
PCV, total red cell count (TRCC) and the haemo-
Prepare a 1/200 dilution of blood in formol- globin (Hb) concentration (g/dl). In the clinical
citrate solution by adding 20 µl of blood to assessment of cases of anaemia it is of value
4 ml of diluent. Wipe the end of the pipette. If to use these indices to calculate the mean cell
a haemocytometer pipette is used, it is neces- volume (MCV), mean cell haemoglobin (MCH)
sary to draw up blood to the 0.5 mark on the and the mean cell haemoglobin concentration
unit, wipe the tip and then draw up diluent to (MCHC).
the 101 mark to make a 1/200 dilution. Allow The MCV indicates the volume of the ‘aver-
2 min for mixing to occur while rotating the age’ red cell in a sample. It is expressed in
pipette. Discard half of the contents and then fill femtolitres (fl; 10 l). Traditionally, MCV was
–15
the chamber. Flush the pipette with the blood/ a calculated parameter, derived by using the fol-
diluent mixture to ensure that all the blood has lowing formula:
been added to the mixing tube. Seal the tube
and invert two or three times to mix well. Fill MCV = (PCV/RBC) × 10
the counting chamber of the haemocytometer
using a fine Pasteur pipette. Make sure that the where RBC is red blood cells. Therefore, if an
counting chamber is just filled and that no fluid animal has a PCV of 42% and a RBC of 6 × 10 /
6
flows into the surrounding channels. Allow the µl = 70 fl.
cells to settle for a few minutes and count using Red cell populations with the MCV below the
the 40× objective. Count the red cells (erythro- reference interval are termed microcytic and those
cytes) in the four corners and the middle square with the MCV above the reference interval are
in the central area, that is, 5 × 16 small squares, termed macrocytic. Automated systems calculate
including those on the right and lower lines and this parameter directly.
excluding those on the left and upper lines (see MCH is the mean cell haemoglobin. This rep-
Figure 5.4). In normal practice a minimum of resents the absolute amount of haemoglobin in
500 cells should be counted (that is, 100 per the average red cell in a sample. Its units are
square). picograms (pg) per cell. The MCH is calculated
from the haemoglobin concentration and the red
cell count using the following equation:
Calculation (depends on the type of
haemocytometer used)
MCH (pg) = [Hb (g/dl)/RBC (× 10 µl)]
9
Total cell counted = N (for example, 500); × 10
volume = 0.2 mm (area counted) ×
2
0.1 mm (depth of chamber) = 0.02 mm 3
Mean cell haemoglobin concentration
Dilution = 1/200 red cell per microlitre = (McHc)
N (500) × 1/0.02 × 200 = 500 × 10,000 =
5 × 10 /µl or 5 × 10 /l MCHC is the mean cell haemoglobin (Hb)
12
6
concentration, expressed in g/dL. It can be
calculated from the Hb and the PCV using the
following formula:

Vet Lab.indb 288 26/03/2019 10:25