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Survival of frozen or vitrified bovine blastocysts produced in vitro in synthetic oviduct fluid



Survival of frozen or vitrified bovine blastocysts produced in vitro in synthetic oviduct fluid



Theriogenology 46(8): 1425-1439



Simplification of in vitro culture conditions offers the advantage of limiting problems due to variation in composition of biological fluids and co-culture with epithelial cells. The aim of this work was to study cryosurvival of bovine embryos produced under partially defined conditions. A conventional freezing technique and a vitrification method that allow for direct transfer of embryos after thawing were compared. Following IVM and IVF, zygotes were cultured in modified synthetic oviduct fluid (SOF), with 10% fetal calf serum (FCS) added 48 h post insemination, or, in the absence of FCS, in a humidified atmosphere of 5%O-2, 5%CO-2, 90%N-2. Morphologically normal Day 8 and 9 blastocysts were classified into 3 groups and were cryopreserved. Group A were blastocysts ( lt 150 mu-m); Group B consisted of expanded blastocysts ( gt 150 mu-m); while Group C were hatching or hatched blastocysts. The freezing solution consisted of 1.36 M glycerol and 0.25 M sucrose in PBS. The vitrification solution consisted of 6.5 M glycerol and 6% bovine serum albumin in PBS (VS3a). Thawed embryos were cultured on granulosa cells and survival was defined as the re-expansion and maintenance of the blastocoel over 24 and 72 h, respectively. Survival rates following vitrification were similar or better than after freezing (Day 8 embryo survival rate at 72 h was 72 vs 57%). More advanced embryos of the same group seemed to survive better. Embryos cultured without serum survived cryopreservation significantly better than those produced with addition of serum at 48 h post insemination (86 vs 61% at 72 h, P lt 0.01). Hatched blastocysts survived freezing or vitrification well (80 to 100%). One calf was born following the transfer of 6 vitrified embryos. The results indicate that both freezing and vitrification methods are suitable for cryopreservation of bovine embryos produced in SOF.

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Accession: 002973138

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DOI: 10.1016/s0093-691x(96)00321-4


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