Procedure
The semen must be killed to prevent movement and diluted before loading into the hemacytometer. This can be done by diluting the semen into a buffer containing a small quantity of formaldehyde. The dilution factor must be recorded to allow calculating the concentration. When there are 20-25 cells per large square (see below) the sample is at the proper dilution.
Loading the Hemacytometer: The tip of the pipette is placed in the V-shaped groove on the hemacytometer to load the sample into the chamber (about 15 microliters.) Capillary action will draw the fluid into the chamber. It is important not to overload the chamber, as doing so will give an inaccurate count. The same is true if the cover slip is moved after the sample is loaded.