The effects of ethylene glycol as a cryoprotective agent on the survival of frozenthawed mouse embryos were examined. The effects of the stage of development of ethylene glycol were also examined.
Eight-cell embryos, morulae, early blastocysts and slid-blastocysts were recovered from superovulated immature ddY mice. Ethylene glycol was added to the embryos in 5 equal increments 5 minutes apart, giving a final concentration of 1.5M. The embryos were cooled to -6℃ at 1℃/min and seeding was induced at -6℃. After being held for a further 5minutes at the seeding temperature, the samples were cooled to -35℃ at 0.3℃/min and then transferred to liquid nitrogen. Rapid thawing was done by placing the straws in 37℃ water. The thawed embryos were diluted in PBS of same time and canners as adding procedures.
Survival of 8-cell embryos and morulae were assessed as a normal development of the embryos to the blastocyst stage and expanding blastocyst after 54 hours and 48 hours of in vitro culture, respectively. While those of the early and mid-blastocysts were assessed to the expanding blastocyst stage after 24 hours of in vitro culture.
The survival rates of 8-cell embryos, morulae, early blastocysts and mid-blastocysts were 73.8%, 74.3%, 87.8% and 77.4%, respectively.
Significant difference on the survival rate among the four stages of development was not observed.
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