Abstract
This study was undertaken to determine the survival of rabbit fertilized eggs after long term storage in liquid nitrogen. Eggs at the 8-16 cell stage were recovered at 45 h post coitum from mature Japanese White rabbits which had been induced to superovulate by treatment with FSH and HCG. The eggs were transferred into 0.1 ml PB1 supplemented with 50% rabbit serum contained in a 1 ml capacity plastic straw. The eggs were equilibrated with 1.5 M DMSO at 37°C for 15 min. The samples were cooled to -76°C at 1°C/min and transferred to liquid nitrogen vapor at -100°C for 5 min. The samples were finally transferred to liquid nitrogen at -196°C and preserved for 210, 4964 and 360463 days, respectively. The samples were warmed from -70°C to -10°C at approximately 4°C/min. At thawing, the DMSO in the medium was diluted in a step-wise manner with freezing medium at 37°C. The eggs were washed with Ringer's solution containing 20% rabbit serum and cultured for 45 days to determine the number of blastocysts developing in vitro. In the other experiments, the frozen-thawed eggs were transferred directly to the oviducts of synchronous pseudopregnant does which allowed to go to term. Statistical significance was determined by the X2 test.
1. The proportions of eggs which appeared morphologically normal after thawing of eggs preserved for 4964 and 360463 days were 82.5 and 71.8%. These figures were not signficantly different from that obtained after thawing of eggs preserved for 210 days, 82.1%. 2. The proportions of eggs developed into blastocysts after culture of frozen-thawed eggs preserved for 4964 and 360463 days were 77.8 and 62.5%. These were not significantly different from that obtained after culture of eggs preserved for 210 days, 63.4%. 3. The proportions of eggs developed into live young after transfer of eggs preserved for 4964 days and 360463 days, 18.5 and 17.9%, were comparable to that obtained after transfer of eggs preserved for 210 days, 16.7%.
