Abstract:

【Objective】 This study was performed to optimize freezing and thawing protocol of rapid boar seminal cryopreservation in 0.5 ml straws. 【Method】 0.5 ml straws were placed at the height of 3, 5, 7, 9 and 11 cm above liquid nitrogen (LN) for 3, 5, 10, 15 and 20 min in polyethylene box, respectively. And the optimal freezing protocol was determined after straws were frozen in LN and thawed in water bath at 37℃ for 30 s. The optimal thawing protocol was determined from six basal protocols, including 37℃ for 30 s, 42℃ for 25 s, 47℃ for 20 s, 52℃ for 15 s, 57℃ for 10 s and 62℃ for 5 s. 【Result】 In twenty-five freezing protocol, both at post-thawed 0 h and 8 h, the percentages of TMS, PMI and NAR of spermatozoa were highest in semen frozen at 3 cm for 10 min. In six thawing protocols, the percentages of TMS, PMI, NAR and Mt-MP of spermatozoa were higher, and the concentration of MDA was lower in semen thawed at higher temperature. Moreover, compared with the control, the percentages of TMS, PMI, NAR and Mt-MP were increased (P＜0.05) and the concentration of MDA was decreased significantly (P＜0.05) in semen thawed at 57℃ for 10 s and 62℃ for 5 s, and there was no difference between thawed at 57℃ for 10s and 62℃ for 5s (P＞0.05). 【Conclusion】 For the rapid freeze-thawing method of this experiment, adopting optimized freezing and thawing protocol increased the quality of thawed semen, meanwhile, freezing 0.5 ml straws at 3 cm for 10 min and thawing at 57℃ for 10 s or 62℃ for 5 s were the optimal protocol.

Key words: optimization, freezing and thawing protocol, rapid freezing, 0.5 ml straw, boar semen