Align-ju STUDENT
Ethylenediaminetetraacetic acid ( EDTA ) sample collection
A high-quality sample is key to obtaining good-quality results . The blood sample should be obtained using the largest-gauge needle appropriate to the patient , and should ideally be a jugular sample if not contraindicated .
The EDTA tube should be filled without the needle attached , and it is important that it is filled to the line . Underfilling the tube can result in excessive anticoagulant . This will lead to the cells shrinking and changes in their morphology . This , in turn , will artificially reduce the mean corpuscular volume ( MCV ) and packed cell volume ( PCV ). If the tube is less than half full , this reduction in PCV can be as much as 5 %. Overfilling the tube will allow clotting to occur and cause a reduction in the number of cells , especially platelets . This may prevent the sample being used for automatic analysis .
The EDTA sample should always be well mixed by inverting it 10 – 15 times once filled , and again before processing in any automatic analyser .
Automated cell counting
The size and shape of each cell is critical to the generation of accurate results by some automated analysers ( quantitative buffy coat , impedance or modified flow cytometer ). This analysis relies on the assumption that erythrocytes are a fixed size . However , cats have variably sized and smaller erythrocytes , and larger , more variably sized platelets , so the analyser can confuse red cells for platelets and vice versa . In addition , in the presence of a disease process that alters the shape and size of erythrocytes , this error can be significantly exaggerated .
The leucocyte count can also be misleading if nucleated erythrocytes are present , for example , in cases of anaemia . The analyser will assume that only white cells are present after lysing the red cells , and will count metarubricytes as mature lymphocytes because , to the analyser , they will appear identical .
blood cell count would be estimated , where necessary , and a check made of whether the counts match the numerical data . The sample would then be examined at × 100 and an assessment made of erythrocyte and leucocyte morphology . The final step could be a semi-quantitative estimation of polychromasia , and a platelet count estimation .
Estimating counts
Platelets Count the number of platelets in 10 highpower fields ( HPF ). Divide the total number by 10 to calculate the average number of platelets per HPF . Multiply the average by 20 to estimate the total count × 10 9 / l .
Leucocytes Estimate at low power (× 10 ) by counting the number of white cells seen in 5 HPF . Calculate the average ; if the count falls between 18 and 50 then the count is likely to be normal . This is a very rudimentary technique , so should be used only as a rough guide .
Erythrocytes These are best calculated by performing a manual PCV , although it can also be useful to look at the thickness of the blood film or the width of the monolayer .
Erythrocyte morphology
Understanding the morphology of erythrocytes can be useful in diagnosing certain disease processes . It is useful to remember a few terms and how they correlate with the automatic cell counts . ( A more detailed glossary of terms can be found at the end of this article .)
Mature erythrocytes are anucleate and appear orangeypink to red . Feline erythrocytes do not show the central pallor of canine erythrocytes , but there is always a mild variation in their relative sizes . Normal erythrocytes are described as normocytic ( normal size ) and normochromic ( normal colour ) ( Figure 1 ).
Blood film examination
The blood film examination should follow a veterinary practice ' s standard operating procedure and should be systematic . An example of a systematic approach would begin with the production of a good-quality blood smear , which is allowed to dry before being stained with an appropriate stain ( such as Wright ' s or Diff-Quik ). The sample would then be examined at × 10 to identify the monolayer . Additionally , the feathered edge would be examined for platelet clumps and large abnormal cells , with a note made of any fibrin strands that may imply micro-clotting , which would alter counts and cell morphology . Next , the white
Figure 1 . Normal red blood cells .
Volume 38 ( 6 ) • December 2023
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