Evaluation of two cryopreservation protocols on cauda epididymal spermatozoa characteristics in domestic cats

Abstract

Various protocols of feline spermatozoa cryopreservation have been reported and used in clinical practice. To compare the quality of post-thawed epididymal spermatozoa subjected to two different sperm freezing protocols, sperm samples were collected from domestic cats (n=7) after castration and divided into 2 aliquots as for Protocol 1 (P1) or Protocol 2 (P2). In P1, after centrifugation, sperm pellets were extended with Tris-based egg yolk extender (TEY) containing 3% glycerol and held for 60 min (5°C) prior to the second dilution (1:1) with 7% glycerol and 1% Equex STM Paste. The straws were frozen by lowering the goblets in three steps into a LN2 tank. In P2, sperm pellets were re-suspended in TEY free of glycerol, left for 15 min at room temperature before the addition (1:1) of TEY containing 8% glycerol and 1% Equex. After loading into 0.25-ml straws at room temperature, the sperm were cooled to 5°C for 25 min and subsequently frozen in a Styrofoam box at 4 cm above LN2 for 10 min. Thawing was done at 37°C for 1 min. No significant differences (P>0.05) between the two protocols (P1 vs P2) in all parameters were observed; total motility (30±8.1 vs 30±3.4; mean±SEM), progressive motility score (3±0.2 vs 3±0.2), percentage of spermatozoa with intact plasma membrane (38.5±4.5 vs 44.5±6.9) and intact acrosome (34±3.2 vs 38±4.7), respectively. In conclusion, both protocols used in the present study yielded similar post-thawed sperm characteristics.