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Beyond the numbers: Things your CBC machine won't tell you, but you need to know (Proceedings)

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Apr 01, 2008

In the diagnostic work up of any case, one of the most frequently used tests is the complete blood count (CBC). The CBC generally includes the gross examination of the sample, the data generated by an automated analyzer and the microscopic evaluation of a blood smear. From the data collected, additional tests may be indicated and/or pursued. Increasingly powerful and easily (for a price of course) available table top hematology analyzers provide an avenue to produce seemingly accurate and precise data rapidly and efficiently. It is very easy to accept the data produced by the analyzer and insert it into an animal's workup. In many instances, one can elect to perform an additional test, choose not to perform a different test, make a diagnosis and begin treatment or change the differential diagnoses list all together. The lure of ease and speed often prove too great for busy clinicians and they use the information from the analyzer exclusively; however, there is a tremendous amount of data that can only be obtained by making a diagnostic blood smear and critically evaluating it using a high quality, diagnostic microscope.

The first step to generating accurate, usable data is to insure the sample was collected and handled in an appropriate manner. A fasting period of 12 hours will generally eliminate the risk of lipemia. Lipemia not only causes artifacts on the serum chemistry panel, but it can also affect the plasma protein, fibrinogen, hemoglobin, mean corpuscular hemoglobin concentration (MCHC) and erythrocyte morphology. A seemingly simple concept to adhere to is sample labeling. In larger practices, with multiple samples being processed by several people, the risk for a handling error increases. Using the appropriate tube is also important. Almost without exception, a vacuum tube with an anticoagulant is used for sample collection. The proper amount of blood must be added to the tube for optimal conditions to be achieved. Glass tubes are less likely to cause clot formation than are plastic tubes. If possible, allow the vacuum of the tube to draw the blood into the chamber, as they are often calibrated to obtain only as much blood as needed, thereby avoiding positive pressure and trauma to the cells. Cell trauma can also be reduced by using a larger bore needle, such as an 18 or 20 gauge to draw blood from a large, free-flowing vein.

For small animals, the anticoagulant of choice is ethylenediaminetetraacetic acid (EDTA), with the hematology tube being the priority tube for filling. It provides optimal retention of in vivo morphology as well as staining characteristics. If there are no contraindications, a liquid anticoagulant is preferred to lyophilized. An incomplete sample draw will decrease the hematocrit (HCT) via cell shrinkage and cause dilution of all cell lines, if liquid anticoagulant is used. Heparin may also be used if WBC clumping occurs. Heparin is not ideal as an anticoagulant for hematologic examinations for multiple reasons. Poor leukocyte staining occurs, likely because heparin binds to them. It is also more prone to platelet clumps forming, and it is not a permanent anticoagulant. A third choice for hematologic sample anticoagulation is citrate.


Figure 1
Once a sample is obtained, it is grossly evaluated within the tube. The tube should be rocked and rolled to check for macroscopically evident clumps or clots. If present, a new sample should be automatically be obtained and no further testing performed. If clinical suspicions suggest the presence of methemoglobinemia, a drop of blood can be placed on white filter paper to see if it is bright red. If the methemoglobin content is >10%, the blood with have a noticeable brown color when placed on white filter paper. If no abnormalities exist, the blood can then be placed into the hematology analyzer and two to four fresh smears can be made as soon as reasonably possible. When the smear is made, additional evaluation for macroagglutination can be performed. A properly made smear should have a head, body, feathered edge and an adequately large monolayer (Fig. 1).

At the same time a blood smear is made, two or more microhematocrit tubes should be prepared and spun in a centrifuge. A packed cell volume (PCV) will be generated to compare to the machine's calculated hematocrit (Hct). Likewise, a crude correlation of the buffy coat can be made to the machine derived white blood cell count (WBC) and platelet count. In addition, the column of plasma can be evaluated for evidence of hemolysis, icterus or lipemia that may affect how all of the data, including the CBC, are interpreted.