COMPARISON OF THE EFFICACY OF CONVENTIONAL SLOW FREEZING AND RAPID CRYOPRESERVATION METHODS FOR BOVINE EMBRYOS

Day 7 bovine morulae and early blastocysts were randomly assigned to o ne of four cryopreservation methods: (i) a modified conventional contr olled slow freezing and stepwise dilution after thawing; and three met hods which enable direct transfer of the embryo into the recipient upo n thawing: (ii) conventional controlled slow freezing and a modificati on of a one-step procedure, (iii) vitrification with 6.5 M glycerol pl us 6% BSA (w/v), and (iv) vitrification with 25% glycerol (v/v) and 25 % propanediol (v/v). In a comparative in vitro study, the percentage o f grade 1 and 2 embryos developing into expanded blastocysts in cultur e for cryopreservation methods 1-4 were, respectively, 53% (29/55), 33 % (20/61), 44% (26/59), and 51% (17/33). Method 2 yielded a significan tly lower survival rate than methods 1 (P < 0.1) and 4 (P < 0.05) and was excluded from a subsequent test of in vivo development. Pregnancy rates (Day 60) after transfer of embryos cryopreserved by methods 1, 3 , and 4 were, respectively, 59% (20/34), 43% (17/40), and 24% (5/21). Method 4 yielded a significantly lower pregnancy rate than method 1 (P < 0.05). Method 3, however, did not yield a statistically different p regnancy rate (P > 0.1) when compared to method 1. Method 3 has consid erable promise in providing a successful method for the cryopreservati on of bovine embryos that (i) reduces the time required for equilibrat ion and cooling, (ii) provides for simple and rapid one-step dilution of cryoprotectant after thawing, and (iii) enables more embryos to be thawed and transferred per unit time. (C) 1995 Academic Press, Inc.