Bautista J A, Dela Peña E C, Katagiri S, Takahashi Y, Kanagawa H
Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
Jpn J Vet Res. 1998 May;46(1):13-8.
Ovulated mouse oocytes denuded of their cumulus cells, were vitrified in a solution containing 7 M ethylene glycol as the sole cryoprotectant using one or two steps of exposure before vitrification and were diluted in 1 M sucrose solution in 5 or 10 min after warming. The results proved that the viability of oocytes are detrimentally affected by exposure to the vitrification solution even without vitrification. At 5 min dilution time, the two-step exposure was superior to the one-step in terms of the post-warming recovery rate of vitrified oocytes with normal morphology and their subsequent development to the blastocyst stage (p < 0.01) after fertilization in vitro. At 10 min dilution time, no significant difference between one- or two-step exposure was found. The effect of the addition of 0.5 M sucrose to the vitrification solution was also determined and did not result in a significant improvement in the viability of oocytes vitrified in one-step and diluted for 10 min. In conclusion, the results in this study indicate that oocytes can be vitrified with 7 M ethylene glycol as the sole cryoprotectant in the vitrification solution, and that the recovery of normal oocytes after one-step exposure in the vitrification solution can be improved by 10 min dilution time. However, the improvement in the recovery rate of oocytes with normal morphology and their subsequent developmental in vitro was not improved by the addition of 0.5 M sucrose to the vitrification solution.
去除卵丘细胞的排卵小鼠卵母细胞，在仅含7M乙二醇作为唯一冷冻保护剂的溶液中进行玻璃化处理，玻璃化处理前采用一步或两步暴露法，解冻后在5或10分钟内于1M蔗糖溶液中稀释。结果证明，即使不进行玻璃化处理，卵母细胞暴露于玻璃化溶液也会对其活力产生不利影响。在5分钟的稀释时间下，就玻璃化卵母细胞正常形态的解冻后恢复率及其体外受精后发育至囊胚阶段的后续情况而言，两步暴露法优于一步暴露法（p<0.01）。在10分钟的稀释时间下，一步和两步暴露法之间未发现显著差异。还测定了在玻璃化溶液中添加0.5M蔗糖的效果，结果并未显著提高一步玻璃化并稀释10分钟的卵母细胞的活力。总之，本研究结果表明，卵母细胞可用7M乙二醇作为玻璃化溶液中的唯一冷冻保护剂进行玻璃化处理，且在玻璃化溶液中一步暴露后，通过10分钟的稀释时间可提高正常卵母细胞的回收率。然而，向玻璃化溶液中添加0.5M蔗糖并未提高正常形态卵母细胞的回收率及其后续体外发育情况。
