Ultrarapid Freezing of Mouse Two-Celi Embryos in Conventional Straws and on Aluminum Foils

Abstract

Objective To compare the success rates of ultrarapid freezing of mouse two-cell embryos in straws and on aluminum foils.

Design               Laboratory experimental study.

Setting               Assisted conception laboratory, Chiang Mai University.

Subjects            Two-cell embryos from superovulated and mated ICR mice.

Intervention      Embryos were divided into 3 groups: 177 were not frozen (control), 172 were

frozen on aluminum foils and 171 in straws. Freezing was done by exposing embryos to 3M DMSO/0.25M sucrose for 2.5 minutes and plunged into liquid nitrogen. Thawing was done by dipping foils into 0.25M sucrose at 37°C or straws into 37°C water-bath and expelled embryos into 0.25M sucrose. Control and frozen/thawed embryos were cultured in G1.2 medium for 24 hours and in G2.2 medium for another 48 hours.

Main Outcome Measures Survival immediately after freezing/thawing, proportions of survived embryos that underwent further cleavage and those that developed into blastocysts and hatching blastocysts.

Results              Embryos frozen on aluminum foils had a significantly higher survival rate than

those frozen in straws (94.8% versus 88.9%; p=.047). They underwent further cleavage at a higher rate (135/163; 82.8% versus 91/152; 59.9%, p<.001). Proportions of embryos that became blastocysts (105/163; 64.4% versus 64/152; 42.1%) and hatching blastocysts (48/163; 29.4% versus 14/152; 9.2%) were also significantly higher (p<.001). Controls had higher (p<.05) cleavage rate (161/177; 91%), and higher (p<.001) rates of blastocyst (136/177; 91%) and hatching blastocyst formation (99/177; 55.9%) than embryos that underwent freezing/thawing.

Conclusion Ultrarapid freezing on aluminum foils gave better results than ultrarapid freezing in conventional straws.