Effect of Different Cryoprotectants on the Survivability and the Development of Bovine Oocytes Matured In vitro

Abstract

Tsuzuki. Y., Kusao. T., Ashizawa, K. and Fujihara, N. 2000. Effect of different cryoprotectants on the survivability and the development of bovine oocytes matured in vitro. J. Appl. Anim. Res., 18: 15–24. To determine the optimum combination of cryoprotectant and freezing method, for improving the developmental, competence of in vitro matured bovine oocytes, three cryoprotectants (1.5 M dimethyl-sulfoxide + 0.25 M sucrose: DMSO; 0.9 M ethylene glycol + 0.8 M propylene glycol + 0.1 M sucrose: EG + PG; 0.8 M PG + 0.8 M DMSO: PG + DMSO) and two freezing methods [the oocytes were cooled from 0C to −6.5C at 1 C/min, seeded and cooled to −30C at 0.3C/min (method A); the oocytes were cooled at −8C directly, seeded and cooled to −35C at 1 C/min (method B)] were used. The survivability and developmental competence of the oocytes thawed from the liquid nitrogen were determined by the dye extrusion assay with eosin and embryonic development after in vitro fertilization, respectively. Without freezing, the survivability of the oocytes equilibrated with and removed from each cryoprotectant were not decreased compared with that in the control group. However, the per cent of the oocyte developed to the blastocyst stage in all of the cryoprotectant treated groups were significantly lower (P<0.05) than that of the control group. DMSO group frozen with method A and EG + PG group frozen with method B maintained their survivability compared to the each control group. However, cleavage rates in all groups were significantly lower (P<0.05)than those of the control groups. No blastocyst was observed in all frozen groups. These results suggest that the combination of DMSO with freezing method A and EG + PG with freezing method B seemed to be more suitable compared with the other combinations.