The impact of lyophilization media on fertilization potential of buffalo spermatozoa

The current investigation aimed to exhibit the impact of lyophilization media of frozen spermatozoa on the fertility potential of buffalo spermatozoa as indicated by comet assay and ICSI. Semen specimens were centrifugated at 700 × g for 20 min using percoll gradient (45-90%), double washed in Tyrode's albumen lactate pyruvate (TALP) and diluted in the lyophilization media (media 1, 2, 3 and 4), correspondingly. Cooling of the diluted sperm cells in vapor of liquid nitrogen. Frozen samples were instantly put into the lyophilizer (-55 °C, pressure 0.001 Mbar). After 24 h of lyophilization, the semen specimens were kept for three months at 4 °C. Frozen-dried semen was re-hydrated at room temperature in of milli-Q water(100 µL). Comet assay results of the frozen-dried semen exhibited that the TCM medium exhibited the lowest % of DNA deterioration [6.17] and the superior % of embryonic developmental rate,while Tris-EDTA medium exhibited the highest % of DNA deterioration [13.09]. The lowest successful % of ICSI exhibited upon using EGTA and EDTA media. It could be concluded that ICSI of frozen-dried spermatozoa upon using TCM medium provides the highest % of embryonic expansion. Also, Tris-EGTA and Tris-EDTA media exhibited the lowest successful percent.