Comparison between Manual and Automated Cryopreservation Techniques and Effects of Different Semen Extenders on Post-Thaw Semen Quality of Buffalo

Cryopreservation has been used extensively for cattle in Bangladesh, albeit no study was conducted on the cryopreservation techniques of buffalo. This study compares two freezing methods and the effects of diluters on the semen quality of buffalo. In the first freezing protocol, semen was frozen in two-step: from 37 °C to 5 °C for 30 minutes in a BLRI-developed equilibration chamber and from 5 oC to -120 oC in a Styrofoam box using liquid nitrogen vapor from different distances (0.5, 1.5, 1.6, 2 and 3 inches). At the same time semen was frozen in a programmable freezer in three steps. The semen samples were then evaluated for motility and morphological quality by CASA. In another experiment, the efficacy of one locally developed diluter-Tris-fructose-egg yolk-based (TFE) and three commercial diluters (Andromed, Triladyl and Steridyl) were evaluated. The highest number of motile sperms (62.67±1.12; P< 0.01) and progressive motility (38.97±1.10; P < 0.001) was observed at 1.6 inches above liquid nitrogen. There was no significant difference in overall motility, progressive and slow motility between semen cryopreserved in the cheap nitrogen vapor technique (57.49±5.67, 38.70± 4.04 and 3.83± 0.63, respectively) and expensive automated technique (65.94±4.65, 45.54 ± 3.64 and 2.43± 0.36, respectively). The highest recovery rate and conception rate were observed in semen diluted with TFE (82.4% and 80%, respectively). Hence, the cryopreservation technique using nitrogen vapor and TFE diluent is cost-effective and suitable for freezing buffalo semen that would produce superior semen for artificial insemination.