Received March 03, 2013
Accepted May 26, 2013

Objectives: 1-Assess the effects of a prolonged period of incubation (8 hrs) prior to vitrification, on ovarian follicle and stroma morphology; 2-Test the effect of low maintenance temperature before ovaries are cryopreservaed; 3-Certify the effectiveness of an aluminium caddy for vitrification of ovarian tissue in terms of primordial and primary follicular morphology and stromal elements after rewarming. Methods: Whole mouse ovaries were left for 8 hrs prior to cryopreservation in HTF medium at 4o.C or room temperature (RT), vitrified inside an aluminium caddy, that was placed into a tightly closed plastic cryovial and then plunged into liquid nitrogen (Lni). Controls were fixed soon after collection. Control and rewarmed ovarian fragments were processed for histology. Slides were analyzed blindly. Results: Primordial follicles presented a superior recovery rate (78%) compared with primary follicles (27%), after being maintained at RT for 8 hrs before vitrification. No significant difference was observed between vitrified and control primordial follicles. When ovaries were maintained at 4o.C, primordial and primary follicle normal morphology rates were 58% and 21%, respectively. Conclusions: Considering that primordial follicles represent the ovarian reserve, these results allow us to suggest that the maintenance of ovarian tissue at RT for up to 8 hours prior to vitrification is a more appropriate procedure for transport or storage than keeping the specimens at 4o.C. In addition, the metal container is an ideal tool for clinical grade ovarian tissue cryopreservation, in fertility preservation programs.