Manual cell counts are performed using a standard hemocytometer, most commonly the improved Neubauer counting chamber as shown in the top image on the right. The Neubauer chamber contains 2 ruled areas, each composed of 9 large squares of equal size. The 9 squares are further divided to facilitate counting larger numbers of cells. A disposable chamber is recommended for CSF samples, especially those from suspected cases of meningitis and Creutzfeldt-Jakob disease. An example of a disposable chamber is the C-Chip, which is a one-piece improved Neubauer hemocytometer with an integrated coverslip. The bottom image on the right illustrates the ruled area of a C-Chip disposable chamber.
Due to the limited precision of manual cell counts, both sides of the hemocytometer should be charged with the fluid and counted. Counts obtained from each side must be within acceptable limits of agreement as established by your laboratory's procedure.
Normally, nucleated cell counts on CSF are quite low, but they can be dramatically elevated in conditions such as meningitis. Specimens that are clear may be counted undiluted if cells do not overlap. Identification of nucleated cells can be improved by rinsing the pipet used to charge the hemocytometer with new methylene blue or crystal violet stain. Phase microscopy also facilitates counting.
Cloudy fluids should be briefly examined microscopically and the necessary dilutions approximated. As fluids are often over-diluted or under-diluted, the laboratory should have clear guidelines concerning the area of the hemocytometer to count and when to make a new dilution.
