Cryotop vitrification of in vitro matured buffalo (Bubalus bubalis) oocytes.

The purpose of this study was to evaluate the efficiency of cryotop vitrification (1) to cryopreserve in vitro matured (IVM) buffalo oocytes, by comparing different concentrations of cryoprotectants. In Group A, in vitro matured oocytes (N=75) were equilibrated in 7.5 % ethylene glycol (EG) and 7.5 % dimethyl sulfoxide (DMSO) for 3 min, transferred in the final vitrification solution (VS), consisting in 16.5 % EG and 16.5 % of DMSO in a base solution (BS) of hepes-buffered TCM199 with 20 % fetal calf serum (FCS) and 0.5 M sucrose, loaded on cryotops, and plunged into liquid nitrogen within 25 sec. In Group B (N=65 oocytes), the equilibration solution was 10 % EG and 10 % DMSO and the VS consisted in 20 % EG and 20 % of DMSO in BS. For warming, oocytes were exposed for 1 min to 1.2 M sucrose and then to decreasing concentrations of the sugar (0.6, 0.4, 0.3 M for 30 sec) in TCM199 + 20 % FCS. Oocytes were rinsed and allocated into the IVM drops for 1.5 h and then fertilized in vitro. The experiment was repeated 4 times. Differences in survival, cleavage and blastocyst rates between groups were analyzed by Chi Square. No significant differences were found between groups A and B in survival rate (92.3 and 92.5 % respectively), cleavage rate (41.7 vs 55.8 % respectively) and blastocyst yield (5.2 vs 8.1 % respectively). It was demonstrated that the cryotop vitrification method is suitable for buffalo oocyte cryopreservation, as indicated by the high percentage of oocytes that survived and cleaved and by blastocyst production. 1) Chian RC, 2004, J Reprod Dev, 50:685-96.