The effects of vitrification on spindle apparatus of in vitro matured germinal vesicle in mice Reza Mahmoudi1 Ph.D., Iraj Amiri2 Ph.D., Parichehr Pasbakhsh3 Ph.D., Iraj Ragardi

Routine oocytes cryopreservation remained an elusive technique in the wide ranges of available assisted reproductive technologies. The microtubules of oocytes are vulnerable to cryoprotectants and thermal change during cryopreservation.

The effects of a vitrification protocol were investigated on the spindle and chromosome configurations of mice oocytes cryopreserved at the germinal vesicle stage.

Germinal vesicle with cumulus cells were transferred to vitrification solution which was composed of 30% (v/v) ethylene glycol, 18% Ficoll-70 and 0.3 M Sucrose either by single step or in step-wise way. Following vitrification and in vitro maturation (MII), the matured oocytes were immonostained for meiotic spindles and chromosomes, before visualization using fluorescent microscopy.

A statistically significant increase was observed in the survival and maturation rate in step-wise vitrification (88.96% and 71.23% respectively) compared to single step vitrification (70.6% and 62.42% respectively) (p<0.05). Normal spindle morphology after vitrification-thawing in step-wise vitrification group (77.26%) was higher than single step vitrification group (64.24%) but lower than control group (94.75%) (p<0.05).

The results suggest that vitrification with step-wise procedure on mice germinal vesicle oocytes has positive effects on survival and maturation rate and normal spindle configuration compare with single step vitrification procedure.