Successful vitrification of day-6 sheep embryos

The aim of the experiments described here was to investigate cryopreservation of day-6 sheep embryos by vitrification methods in which the preliminary procedures can be performed at room temperature using VS1 (5.5 mol ethylene glycol l-1 and 2.5 mol glycerol l-1), VS11 (6.0 mol ethylene glycol l-1 and 1.8 mol glycerol l-1) and VS14 (5.5 mol ethylene glycol l-1 and 1.0 mol sucrose l-1). None of the day-6 sheep embryos vitrified with VS1 survived. Day-6 sheep embryos with the exception of blastocysts were vitrified with VS11 with no loss of viability in vitro. The viability of transferred day-6 embryos vitrified with VS11 was however extremely poor. Osmotic damage was avoided by initially exposing the embryos to one of four dilutions (20%, 30%, 40% and 50%) of VS11 for 5 min at 25 degrees C and then vitrifying with the undiluted VS11. The highest survival (88.2%) in vitro was obtained when embryos were exposed to 30% VS11 before vitrification with the undiluted VS11. Survival of transferred embryos exposed to 30% VS11 and then vitrified with undiluted VS11 was 55% (16 of 29) for morulae and 62% (18 of 29) for blastocysts. The pregnancy rate for recipients that received two vitrified sheep embryos of these developmental stages per ewe was 79% (22 of 28). In a small study performed with VS14 the survival of day-6 sheep embryos vitrified with VS14 (in one-step) was 100% in vitro and 50% after transfer.