Cryopreservation of bovine blastocysts obtained by intracytoplasmic sperm injection

The freezability and survivability of zona-intact and zona-free (hatched) b ovine blastocysts obtained by intracytoplasmic sperm injection (ICSI) were assessed. Day 7 or 8 blastocysts were cryopreserved by slow freezing using 1.5 M glycerol and 0.2 M sucrose. Embryos were exposed to solutions in a 2- step procedure at room temperature and frozen in a programmed cell freezer. Blastocysts that re-expanded within 6 h of post-thaw culture were consider ed viable. The cleavage, morula and blastocyst development rates after ICSI were 52.4 (131/250), 39.7 (52/131), and 24.4% (32/131), respectively. Blas tocyst stage embryos were randomly divided into 2 groups. The first group o f embryos was frozen with their zonae intact, while the second group was al lowed to hatch from their zonae during the additional 18 h culture, after w hich they were frozen. The data showed that more Group 2 blastocysts (14/16 , 87.5%) than Group 1 (12/16; 75.0%; P<0.05) survived, and more zona-free b ovine blastocysts frozen with glycerol as the cryoprotective agent (CPA) th an zona-intact blastocysts after slow freezing retained their viability. (C ) 2000 by Elsevier Science Inc.