title: Vitrification of bovine blastocysts obtained by in vitro culture of oocytes matured and fertilized in vitro.
abstract: Two experiments were conducted to assess the viability of bovine blastocysts obtained by in vitro fertilization of oocytes matured in vitro (IVM-IVF) and cryopreserved by vitrification. In Expt 1, the optimal concentrations of glycerol and 1,2-propanediol in the basic medium (modified TCM199) for cooling and warming without formation of ice crystals were determined by plunging the solution into liquid nitrogen and then warming it in a water bath at 15 degrees C; when both glycerol and 1,2-propanediol were present in the solution (> 45% v/v), vitrification of the medium was observed. In Expt 2, IVM-IVF blastocysts were equilibrated to the mixture of glycerol and 1,2-propanediol (0% to 45%) at 15 degrees C in a stepwise manner as follows: (i) in one step, for 18 min to the final vitrification solution; (ii) in two steps, for 8 min in the first step and 10 min in the second step; (iii) in four steps, for 4 min in the first three steps and 6 min in the last step; (iv) in eight steps, for 2 min in each step, but 4 min in the last step; and (v) in 16 steps, for 1 min in each step, but 3 min in the last step. After removal of cryoprotectants, the blastocysts were cultured for 24 h in vitro. The survival rates for the embryos equilibrated in 1, 2, 4, 8 and 16 step(s) were 56, 89, 100, 100 and 100%, respectively. The blastocysts equilibrated in 1, 2, 4, 8 and 16 steps were vitrified by plunging the straws containing them into liquid N2, thawed and cultured in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Tokyo Bio-Technology Center, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Kuwayama
authors.forename: S
authors.initials: S
authors.lastname: Hamano
authors.forename: T
authors.initials: T
authors.lastname: Nagai
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000166:cytology; D002417:Cattle; D002478:Cells, Cultured; D015925:Cryopreservation / Q000379:methods*; D004624:Embryo Transfer / Q000379:methods; D005307:Fertilization in Vitro; D005328:Fetal Viability; D009866:Oogenesis