White Blood Cell Count (WBC)
The white cell count (WBC) is the total number of leukocytes in a volume
of blood, expressed as thousands/Ál. As with the RBC, the WBC can be
done by manual methods or by automated cell counters. The WBC by any
method is a count of nuclei or total nucleated cell count. If nucleated
red blood cells (nRBC) are circulating in blood, they will be included in the nucleated
cell count whether the count is done by manual methods or by automated
analyzers. In such cases, the WBC represents the leukocyte count only
after it has been corrected for the nucleated red cells (nRBCs). The
number of nRBCs per 100 leukocytes is recorded during the differential
leukocyte count. Then a correction is made as follows:
Corrected WBC = nucleated cell count x (100
÷ [nRBC + 100])
In reports from our laboratory (excluding automated hematology
tests), the WBC value is always a leukocyte count since we correct for the presence of nRBCs if > 5 nRBCs are counted in a 100 cell differential leukocyte count.
Manual WBC counting
The manual procedure, using a diluting chamber and a hemocytometer, is
no longer used except for white cell counts in body fluids with extremely
few cells (< 1,000 cells/µL), e.g. cerebrospinal fluid. It involves
diluting blood in a diluent that lyses
the red cells to remove them from view. A
hemocytometer is charged with the diluted blood, and nuclei are counted
in the appropriate areas of the grid using a light microscope. The manual
WBC is a reasonable test to do in-office since it is not terribly time-
and labor-intensive and gives acceptably accurate results.
Laboratories use either impedance-based electronic cell counters (see
RBC section) to generate WBC counts in body fluids and non-EDTA samples.
In our laboratory, the Coulter Z2 (a bench-top impedance counter) is
our back-up analyzer. We use this cell counter when our Advia is out
of commission or a sample other than EDTA-anticoagulated blood is submitted
for cell counts. This is the standard instrument used for cell counts
in body fluids, other than blood, e.g. joint fluids, peritoneal fluids,
because these fluids cannot be analyzed through the Advia.
Flow cytometry counters
We use the hematology analyzer, the Advia, to obtain WBC counts. This analyzer counts WBC in 2 ways, both of which use flow cytometry.
In the first method called the peroxidase method, the white cells
are stained with peroxidase and the cells are counted based on size
and staining characteristics.
E. Large unstained cells
This method also provides an automated differential cell count by separating
the cells into clusters (see image on left for a dog peroxidase cytogram).
The second method, called the basophil method, involves stripping
the cells of cytoplasm and counting nuclei (see image below). In this
cytogram, the cells are displayed as a worm, with the mononuclear cells
(lymphocytes and monocytes) in the head and the granulocytes
(neutrophils) in the body of the worm. This is a far more accurate method
for counting white cells and is the default method on the Advia. The
peroxidase method will give falsely high counts in samples with
numerous platelet clumps. This type of artifact is also prominent with impedance-based counters, resulting in erroneously high WBC counts in certain
species, especially cats (whose platelets clump at the drop of a hat).
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