Open pulled straw vitrification and slow freezing of sheep IVF embryos using different cryoprotectants

Abstract

The aim of the present study was to evaluate the post-thaw survival and hatching rates of sheep blastocysts using different cryoprotectants. In Experiment 1, Day 6 sheep embryos were cryopreserved by a slow freezing protocol using 10% ethylene glycol (EG), 10% dimethyl sulfoxide (DMSO) or a mixture of 5% EG and 5% DMSO. Hatching rates were higher in the 10% EG group than in the 10% DMSO or EG + DMSO groups (30% vs 18% and 20%, respectively). In Experiment 2, embryos were cryopreserved by open pulled straw (OPS) vitrification using either 33% EG, 33% DMSO or a mixture of 16.5% EG + 16.5% DMSO. Re-expansion and hatching rates in the EG + DMSO group (79.16% and 52.74%, respectively) were higher than those in the EG group (64.28% and 30.02%, respectively), whereas the outcomes for the DMSO group were the lowest (45.18% and 8.6%, respectively). In Experiment 3, embryos were cryopreserved by OPS vitrification using either 40% EG, 40% DMSO or a mixture of 20% EG + 20% DMSO. Re-expansion and hatching rates were highest in the EG group than in the EG + DMSO and DMSO groups (92.16% vs 76.30% and 55.84% re-expansion, respectively; and 65.78% vs 45.55% and 14.46% hatching, respectively). In conclusion, OPS vitrification was found to be more efficient for cryopreservation of in vitro-developed sheep embryos than traditional freezing.

Cryopreservation and Transfer of Sheep Embryos Recovered at Different Stages of Development and Cryopreserved Using Different Techniques

“…Thus, the results of the present study, using both traditionally accepted slow freezing and ultra-fast vitrification for embryo cryopreservation show that embryo quality was good with both techniques. The results of our study are consistent with those obtained by other researchers [45,46], in which the quality of frozen-thawed embryos was higher with vitrification procedures than with the slow-freeze method [47,48]. The advantages of using vitrification for embryo cryopreservation are simplicity, rapidity of the procedure and the fact that this method does not require expensive equipment as in the case of the slow freezing method.…”

Resveratrol Reduces ROS by Increasing GSH in Vitrified Sheep Embryos

“…Following Bhat et al [15], 10% of the best-quality early blastocysts of each experimental group were vitrified using the open pulled straw (OPS) technique [16], which consisted of washing the blastocysts in 800 µL of holding medium (TCM-199 without HEPES, supplemented with 10% FBS). They were then sequentially equilibrated, first in 600 µL of holding medium supplemented with 7.5% of ethylene glycol and 7.5% of DMSO for 3 min, then in 600 µL of holding medium with 0.342 g/mL of sucrose, 16.5% of ethylene glycol, and 16.5% of DMSO for 20 s. Immediately afterwards, 2 µL drops were formed with the embryos and collected by capillarity in an OPS.…”

Aquaporins and Animal Gamete Cryopreservation: Advances and Future Challenges

“…Caprine embryos have been shown to present better viability after the vitrification process than after controlled-rate slow freezing [73][74][75]. On the other hand, studies performed in equine and donkey embryos showed that controlled-rate slow freezing seemed to be the most appropriate cryopreservation method [76,77].…”