Optimized Method for Bovine Blastocyst Vitrification Using a Simple Hand-Made Cryotip

Abstract

Objective: This study introduced a simple method for bovine blastocyst vitrification.Materials and Methods: Bovine blastocysts were produced in vitro by means of a wholeco-culture system with vero cells. The blastocysts were randomly divided 1:3 into either vitrification(100 blastocysts) or control (43 blastocysts) groups. For vitrification,expanded - blastocystswere incubated first in equilibration medium for 8 minutes and then in the vitrificationsolution for 1 minute. The blastocysts were then loaded in the tip of a handmade cryotip forimmediate - deep freezing in liquid nitrogen. Frozen embryos were then warmed by directlyimmersing the tips in sequential warming solutions . warmed embryos were cultured for afurther period of 48 hours when the ratios of re-expansion, hatching and degeneration werecompared with the control group.Results: After warming, in the vitrified and control groups the ratios of re-expansion were78.5% ± 0.067 and 81.6% ± 0.072, the ratios of hatching were 43.7% ± 0.083 and 49.8%± 0.089 and the ratios of degeneration were 36% ± 0.082 and 22.3% ± 0.087, respectively,which were not significantly different between the two groups.Conclusion: Post - warming survival of the vitrified and non - vitrified embryos were notsignificantly different, handmade cryotips can be used as an efficient and feasible device forbovine blastocysts vitrification.

Keywords

• Blastocyst
• Warming
• Survival