In order to cryopreserve germinal vesicle (GV) stage oocytes, we first need to develop a novel container for keeping large quantities of GV oocytes, because of collecting them as cumulus oocytes complexes (COCs) that have bigger size and larger volume than oocytes themselves, and second modify a protocol for optimizing vitrification of them. In this mini-review, we describe our recent progress for attaining these objectives. When 65 bovine COCs having GV oocytes could be placed on a sheet of nylon mesh, and plunged directly into liquid nitrogen for vitrification, the recovery rate was significantly higher compared with that in 15 ones on the electron microscope (EM) grid as a control, followed by obtaining the resultant cleavage and developmental rates after in vitro fertilization and culture (IVFC) without significant difference. Using bovine and murine oocytes, we found that a step-wise manner to expose them with the vitrification solution increased rates of in vitro maturation, subsequent development to blastocysts and hatching/hatched blastocysts after IVFC. Our results show that nylon mesh is an alternative material for cryopreserving large quantities of bovine GV oocytes, and that a step-wise exposure to cryoprotectants may have befit for decreasing disadvantage during vitrification
