High survival of mouse oocytes using an optimized vitrification protocol.

Abstract

The method of vitrification has been widely used for cryopreservation. However, the effectiveness of this method for mammalian oocytes could be improved by optimizing each step of the process. In the present study, we tested the effects of varying several key parameters to determine the most effective protocol for mouse oocyte vitrification. We found that cryoprotectant containing ethylene glycol and dimethylsulfoxide plus 20% fetal calf serum produced the highest rates of oocyte survival, fertilization, and blastocyst formation. The duration and temperature of oocyte exposure to vitrification and thawing solutions influenced survival rate. The presence of cumulus cells surrounding oocytes and the incubation of thawed oocytes in Toyoda-Yokoyama-Hosoki medium also increased oocyte survival. Open pulled straw and nylon loop methods were more effective than the mini-drop method. Finally, the combination of these improved methods resulted in better spindle morphology when compared to the unimproved methods. These results demonstrate that the outcomes of mouse oocyte vitrification can be improved by a suitable combination of cryopreservation methods, which could be applied to future clinical research with human oocytes.