New perspectives in laboratory quality control of canine semen processing.

A cryopreserved insemination dose is usually calculated by measurement of concentration and morphology of spermatozoa in pre-processed semen and the motility in thawed semen. Accurate measurement of spermatozoal concentration enables calculation of the insemination dose of spermatozoa; however, all methods used for measuring spermatozoal concentration have inherent measurement errors and therefore proper laboratory quality control is essential. In our laboratory fluorescent microscopy is used for counting and live-dead assessment. Our data show often-substantial discrepancies in the calculated spermatozoal concentration extrapolated from measurements in neat semen to the concentration measured after thawing. Similar variability was observed in samples cryopreserved by external centers using extrapolated calculations. Computer-assisted semen analysis (CASA) often overestimated semen concentration in our laboratory yet is frequently used when processing semen. The method employed when processing is often not recorded. Because spermatozoal concentration data from external centers may be unreliable, full post-thaw semen analysis is often required at the time of insemination to adjust the dosage appropriately. This extra analysis adds unnecessary time and complexity. Assessment of motility is usually not so problematic. Morphological evaluation of spermatozoa is important, but detailed reporting of defects is often incomplete and may rely on estimation rather than direct counting. The commonly used measure of progressively motile normal spermatozoa (PMNS) may therefore be inaccurate. Recommendations are provided for canine semen cryopreservation, including routine direct measurement of spermatozoal concentration in thawed semen.