Efficacy of Histoprep® gradient for isolating ovine epididymal sperm

The aim of the present study was to compare Histoprep® continuous gradient and swim-up procedures for the isolation of ram epididymal sperm. The ram testes-epididymides were collected from the local slaughterhouse and stored in a refrigerator (4-6 °C) for 24 h. On the day of in vitro fertilization (IVF) cauda-epididymides were separated, incised, and incubated in warmed Bracket and Oliphant medium (BO; 2 mL) for 15 min. The swim-up procedure was performed by the addition of 800 μL of BO to sperm suspension (200 μL) and incubation in a CO2 incubator for 30-45 min. The same volume of sperm was overlaid on Histoprep® and the company's instructions were followed for the isolation of human lymphocytes. Sperm parameters were analyzed before and after isolation. Isolated sperm was subjected to IVF with in vitro matured ovine oocytes. The proportion of progressive motile sperm were 1.14 ± 0.03 and 1.22 ± 0.05 after the Histoprep® and swim-up procedures, respectively (P > 0.05). The proportion of isolated sperm by Histoprep® (0.1 ± 0.01) was significantly higher than the swim-up (0.06 ± 0.01) procedure (P < 0.05). The efficacy of the Histoprep® and swim-up procedures for sheep cleavage (62 ± 5.21 and 64.4 ± 12.35%) and blastocyst (6.8 ± 1.1 and 7.9 ± 1.21%) rates was similar (P > 0.05). The results of the present study showed comparable efficacy of the Histoprep® continuous gradient for ovine epididymal sperm isolation with the swim-up procedure in terms of sperm quality and fertilization output, with a higher recovery rate.