USEFULNESS OF POLYETHYLENE-GLYCOL FOR CRYOPRESERVATION BY VITRIFICATION OF IN VITRO-DERIVED BOVINE BLASTOCYSTS

A series of five experiments measured the high survival of bovine blas tocysts produced in vitro after cryopreservation by vitrification. The vitrification solution (designated VS) contained 40% (v/v) ethylene g lycol, 6% (w/v) polyethylene glycol and 0.5 M sucrose in phosphate-buf fered saline. Embryos developed in vitro at Days 7 and 8 (Day 0 = inse mination day) were exposed in one step to VS for 1 min or two steps wi th 10% ethylene glycol for 5 min and then VS for 1 min. In both cases, the embryos were finally cryopreserved in liquid nitrogen. After the embryos were warmed rapidly and the VS solution diluted, the survival rates were assessed by monitoring hatching rate in vitro. They were 13 .0% for the one-step and 72.7% for the two-step procedures (P < 0.001) . When embryos were exposed to individual solutions containing 6% (w/v ) of each of 4 macromolecules (polyethylene glycol, BSA, polyvinylpyrr olidone or Ficoll) in the two-step protocol and then cryopreserved, th e survival rates were 79.3, 34.8, 41.4 and 57.1%, respectively. After embryos had been exposed to the VS in two steps and then cryopreserved , there were no significant differences in survival rates when the sol utions were diluted with or without sucrose. These results indicated t hat a vitrification solution containing polyethylene glycol can be use d for cryopreservation of bovine blastocysts produced in vitro, and th at a two-step addition of VS improved the in vitro survival of post-wa rming embryos. It was also shown to be possible to dilute post-warming embryos directly without the use of sucrose solution. (C) 1997 Elsevi er Science B.V.