Vitrification of Mouse 4-Cell Embryos by OPS Method

The experiments were conducted at 37℃ to cryopreserve mouse 4-cell embryos by vitrification with EFS30,EFS40,EDFS30 and EDFS40(an ethylene glycol-based vitrification solution) respectively in open pulled straw(OPS).In the 1-step OPS method,embryos were vitrified after direct exposure to vitrification solutions for 15s,25s or 35s,while in the 2-step method,the superovulated embryos were first pretreated to either 10% Ethylene glycol(EG) or 10%EG+10%DMSO for 30s respectively,then exposed to a vitrification solution(VS) for 25s,30s or 35s and then immersed in liquid nitrogen.After warming,the survival of embryos was assessed by their ability to develop to blastocysts or develop to term after transfer.The higher blastocyst rates of vitrified embryos were from EDFS30 group in the 2-step OPS method(88.6%) and EDFS40 group in the 1-step method(87.7%),which were similar (P0.05) to that(93.0%) obtained in non-vitrified fresh embryos.The embryos derived from the best vitrified group or fresh 4-cell embryos(used as control) after culture for 1 to 3 h were transferred to 50～60 h pseudo-pregnant female mice.17 fetuses were obtained from 4 of 12 recipients which had received 143 vitrified-warmed embryos.In the pregnant recipients,the percentage of transferred embryos developed to young in the treated group(42.5%) and control(59.4%) showed no difference (P0.05).These results demonstrated that mouse 4-cell embryos could survive cryopreservation by vitrification in OPS.