The primary objective of the semen processing laboratory is the production of high quality artificial insemination doses. High quality equates to normal, motile cells extended to a known concentration.

The hemacytometer is commonly used to quantitatively measure sperm concentration in ejaculates. Freund and Carol (1964) described sources of variation, including technician, preparation and number of dilutions and chambers, and counting error.  To date, the hemacytometer has been considered the ‘gold standard’ for measurement of sperm concentration, probably since it represents actual cell counts instead of via a mechanical method, even though error can be high.

The electronic photometer is a useful  technique to measure sperm concentration in ejaculates. Calibrations are based upon correlations of  %T (O.D.) typically using hemacytometer counts of multiple ejaculates ( Foote, 1968; 1972). Photometers using incandescent light sources must be calibrated on a routine basis, especially if the light source is replaced. The recent use of LED lamps eliminates this requirement. Filters or  standards are often used to confirm machine status. Technician, non-sperm cells or debris, light reflection/refraction, dilution preparation and machine contribute to variance.

CASA systems ( Amann and Waberski, 2014, review) have more recently been used to automate sperm concentration measurement. Error is associated with system, technician, software set-up, dilutions and chamber preparation. Amann and Waberski note that, at least with present technology, CASA systems may not provide highly accurate and precise concentration measurements.

The NucleoCounter SP100 (ChemoMetec, Denmark) has most recently been validated (Hanson et al, 2006) as a useful tool for measurement of sperm concentration in neat or extended samples. There is no interference from extenders, lubricants, non-sperm cells, etc. Variance is primarily associated with sample preparation (pipetting). Expected CV’s are 5-6%. Due to its accuracy and repeatability, it is suggested that the SP100 be now considered the ‘gold standard’ technique.