Simple storage (CO2-free) of porcine morulae for up to three days maintains the in vitro viability and developmental competence

Abstract

The advancement of porcine embryo transfer (ET) technology is constrained by regulatory hurdles (liquid nitrogen transportation) or, more importantly, the technical obstacles of using vitrified embryos in combination with nonsurgical deep uterine ET technology. Maintaining embryos in culture during transport and prior ET collides with the need of CO₂ gassing and the best choice of culture medium. In this work, we describe storage conditions for short-term embryo CO₂-free storage that allowed for a majority of in vivo-derived porcine morulae to survive after 3 days of storage in a liquid state, and to develop to the blastocyst stage unhatched, a sanitary prerequisite for ET. The storage conditions included NCSU-23 medium supplemented with bovine serum albumin, where bicarbonate was partially replaced by HEPES to avoid the need for CO₂ gassing, and a temperature of 37 °C. These conditions were able to maintain the functionality of the stored embryos (hatching capacity after exposure to conventional culture conditions) and their developmental competence after ET (normal fetuses by day 38 of pregnancy). Use of this strategy of CO₂-free storage should allow the shipment of fresh embryos worldwide in the absence of liquid nitrogen.

Keywords: Blastocyst; CO(2)-free embryo storage; Embryo transfer; Morulae; Pig.