116 Evaluation of frozen–thawed bovine embryos before and after 24-hour culture

Fertility is a lowly heritable trait that has an exceedingly large financial impact and may limit the ability to make genetic progress. Acceptable pregnancy rates using assisted reproductive techniques (ART) such as IVF and cryopreservation, relies upon having accurate tools. Embryo cryopreservation is an enabling ART for cattle producers and embryo technicians alike, that allow producers to reach financial and production goals. There has been a shift from post-thaw processing to freezing embryos in ethylene glycol (EG) to allow post-thaw direct transfer to recipients. Due to this, we have little information about the post-thaw viability or morphology of bovine embryos frozen for direct transfer. The objective of this research was to evaluate whether there is cryoprotectant dependent effect(s) upon embryos post-thaw, which translates to suppressed developmental capacity in the EG-treated embryos. In vivo-derived embryos frozen in either EG (n = 48) or glycerol (GLY; n = 48) were evaluated for grade and stage post-thaw (International Embryo Technology Society). Pre-freeze data regarding grade, stage, and donor was not made available for these donated embryos. On average, thawed EG embryos were at an advanced stage at ~7 d when compared to GLY embryos. Embryos were rinsed and relocated into 5-μL drops of culture media covered in oil for 24 h at 5% CO₂, 5% O, 90% N, and 100% humidity. Following culture, grade and stage were once again assessed. Embryos that continued to develop in culture were deemed viable and the developmental grades and stages were analysed with ANOVA, while the percent that continued was analysed with chi square. Culture was used to estimate viability post-thaw, and those embryos that continued to develop were deemed viable. A table outlining this information would not fit, and therefore the information has been summarised in the following text. Although the EG-treated embryos were at an advanced (P < 0.05) stage relative to the GLY treated embryos post-thaw (4.8 ± 0.2; 4.1 ± 0.1, respectively), the GLY embryos advanced to a more (P < 0.05) developed stage during culture than EG embryos (5.9 ± 0.2; 5.3 ± 0.2, respectively). As a group the EG embryos advanced 0.5 developmental stages, or an 11.7% change, compared to an advancement of 1.8 developmental stages in the GLY embryos, a 44.4% change. The post-thaw to postculture grade of the EG-treated embryos tended (P < 0.1) to decrease (1.6 ± 0.1 to 1.9 ± 0.2) while the grade of the GLY treated embryos did not change statistically (1.8 ± 0.1 to 1.9 ± 0.2). Cryoprotectant dependent effect(s) upon the embryo viability post-thaw translated to suppressed developmental capacity in the EG-treated embryos. Although convenient for the ET practitioner, embryos frozen in EG may suffer substantial damage that is currently unevaluated. Future research will involve pre-freeze and post-thaw evaluations and perhaps transfer of cultured embryos into recipients to determine whether freeze/thaw damage can be overcome with a 24-h in vitro culture period.