[Influence of cryoprotectant and cooling rate in vitrification method on the spindles of rabbit oocytes].

OBJECTIVE

To investigate the influence of cryoprotectants and cooling rates in vitrification method on the spindles of rabbit M II oocytes.

METHODS

Rabbit oocytes were verified by using cryoloop with ethylene glycol (EG) singly or EG combined with dimethyl sulphoxide (DMSO) as cryoprotectants, and cooled by taking oocytes directly into liquid nitrogen or by vitrification machine. After frozen rabbit oocytes thawed, the microtubulin and chromosome of the spindles were fixation and stained by immunofluorescent method. Confocal microscope was used to reveal spindle configuration.

RESULTS

In the two protocols of single EG used and EG combined with DMSO, the spindles were severely injured. But in protocol of EG combined with DMSO and at ultra-rapid cooling rate, the normal configuration of spindle rate of thawed rabbit oocytes was similar to that of the control group.

CONCLUSION

The protocol of EG combined with DMSO as cryoprotectants and with extremely high cooling rate by vitrification machine can produce the best effect on conservation of spindle configuration in vitrification of rabbit oocytes.