Manual collapse of blastocoels in not effective in increasing the viability of vitrified equine embryos

Abstract

Embryo cryopreservation methods have been used for commercialization and formation of genetic banks. Cryopreservation of equine embryos <300 µm in diameter, collected at days 6-6.5 after ovulation, allows satisfactory pregnancy rates. However, higher embryo collection rates in mares are obtained when uterine flush is performed between days 7 and 8 after ovulation when embryos are >300 µm in diameter, needing blastocoel collapse for satisfactory resistance to cryopreservation by vitrification. To evaluate the viability of simplified blastocoel collapse by embryo puncture with low technology and low-cost equipment, 22 embryos, collected at day  8 post-ovulation (D8), were allocated to the following groups: (1) micropuncture with a 30 G needle, assisted by a mechanical micromanipulator, before vitrification (n=4); (2) manual blade microsection before vitrification (n=6); (3) no manipulation prior to vitrification (n=8); and (4) freshly inovulated embryos (n=4). Despite the high re-expansion rates observed after vitrification, embryos manipulated prior to vitrification (groups MP and MS) did not result in pregnancy 25 days after transfer. On the other hand, embryos from groups NM (non-micromanipulated) and FR (freshly inovulated) resulted in pregnancies at 25 days. Under the conditions of the present study, manual blastocoel collapse was not efficient in increasing cryotolerance to vitrification among large embryos, requiring improvements to obtain pregnancies.