Cryoprotection of Day-4 mouse embryos by methanol

Methanol was examined as a cryoprotective additive that permits the direct transfer of frozen--thawed Day-4 mouse embryos to foster mothers without dilution of the cryoprotectant. Methanol permeated the embryos rapidly, was not toxic and exerted a cryoprotective action. The highest level of survival (50%) of embryos in vitro was observed after equilibration in Medium PB1 containing 3.0 M-methanol, slow cooling (0.5 degrees C/min) to a temperature between -30 and -40 degrees C, rapid cooling (800 degrees C/min) and storage in liquid nitrogen (-196 degrees C), rapid warming (800 degrees C/min), and rapid dilution. A high rate of development in vivo to late-stage fetuses (up to 81%) was observed when cryopreserved embryos were transferred to pseudopregnant recipients immediately after thawing.