大鼠早期胚的快速冷冻保存

Frozen Storage of Rat Embryos by Quick Freezing Method

为建立大鼠胚胎冷冻保存库探索一种经挤有效的冷冻处理技术,本文着重探讨了两种不同的快速冷冻处理方法(1.把胚胎先在—30℃下保持60分钟后投入液氮;2.在液氮蒸气中停留5分钟后投入液氮)及四种不同的防冻液(添加2.8M甘油,0.5M蔗糖的PBS+20％大鼠血清或胎牛血清,生理盐水+20％大鼠血清或胎牛血清)对大鼠早期胚冷冻-解冻后发育率的影响。实验结果表明,在液氮蒸气中保持5分钟后投入液氮比在-30℃下保持60分钟的效果好,防冻液内添加胎牛血清(FCS)比大鼠血清(RS)效果好,PBS与生理盐水之间无明显差异。以含有2.8M甘油,0.5M蔗糖的PBS+20％FCS作为防冻液,用第二种方法进行冷冻处理的大鼠早期胚解冻后其发育率为81％,把用上述方法处理的135枚发育胚移植到17只受体鼠的子宫角内,有2只妊娠产仔4只。

In order to find a economic and efficient method for establishing the bank of frozen rat embryos, the two different quick freezing methods and four kinds of freezing media were evaluated in this study. In method A the straws were inserted into the—30℃ chember of computer controled embryo—freezer for 60 min., then plunged into LN_2and in method B the straws were held vertically in the neck of LN container for 5 min., then plunged into LN_2derectly. The composition of four freezing media were PBS and Saline (0.9%NaCl) supplement with 20% rat serum and fetal calf serum respectively. 2.8M glycerol and 0.5M sucrose were used as cryoprotectant added in freezing media above. The results showed that method B was better than method A(P<0.05). The development rate of embryos frozen with method Breached 81.0%. There were no differences between PBS and saline, but FCS was apparently better than rat serum (p<0.05). The 135 frozen—thawed and developed embryos were transferred into 17 recipients and 2 became pregnant with 4 baby rats.