Canine Sperm Concentration: Comparison Between Neubauer Cell Counting, Photometers and a CASA System

This study aimed to compare three methods used in evaluating canine sperm concentration in translucent and opaque media. These techniques were counting chamber; spectrophotometry using two commercial photometers calibrated for canine semen (Accuread and SDM1); and CASA analyzer (Hamilton Thorne IVOS II [HT-IVOS II]), for motile and non-motile spermatozoa (spz). Eight ejaculates were collected, then the sperm-rich fraction of each sample was divided into two aliquots: one was mixed with a translucent commercial buffered solution (Easy Buffer-B) and the other with an opaque egg yolk-based diluent. Each of the two aliquots was separated into five fractions evaluated respectively by the different tools. The results showed that the HT-IVOS II analyzer with immobilised spermatozoa was in good agreement with the reference method (counting chamber), whereas the Accuread photometer showed a reasonable difference, but with poor statistical agreement, while the SDM1 photometer and HT-IVOS II analyzer (with immobilised spz) were in bad agreement with the reference method in assessing translucent media-diluted canine sperm concentration. In the context of semen diluted in an opaque egg yolk-based medium, none of the studied techniques was in agreement with the reference method. The comparison between translucent media-diluted and opaque media-diluted semen revealed that except for the reference method, all evaluation techniques (SDM1, Accuread and HT-IVOS II analyzer) were unreliable (p > 0.05) in assessing sperm concentration across both situations. The authors propose to conduct further studies with an expanded sample size and to incorporate flow cytometry (FCM) into the comparative evaluation techniques.