IMPROVED YIELDS OF BOVINE BLASTOCYSTS FROM IN-VITRO PRODUCED OOCYTES .1. SELECTION OF OOCYTES AND ZYGOTES

Three experiments were conducted with 7,901 oocytes (6,152 cleaved zygotes) to identify methods to increase the proportion of oocytes aspirated from ovaries of slaughtered cows that would develop to expanded blastocysts by 9 d after exposure to spermatozoa. In the first experiment, oocytes were matured for 24 h in TCM-199, fertilized, and presumptive zygotes were co-cultured in TCM-199 and granulosa cells. Cleaved zygotes from oocytes classified as good on the basis of quality of investments and cytoplasm developed to expanded blastocysts more frequently than did zygotes from intermediate-marginal oocytes (29% vs 24%; P=0.035). In the second experiment, marginal oocytes were subdivided into those with partially expanded cumulus, coarsely granulated cytoplasm or minimal cumulus (not more than 3 layers of cumulus cells, or zona pellucida partly exposed), matured in Ham's F-10, and co-cultured in Menezo's B2 with buffalo rat liver (BRL) cells. Cleaved zygotes from oocytes with partially expanded cumulus and/or coarsely granulated cytoplasm developed to expanded blastocysts as frequently (47%) as did good/intermediate oocytes (40%), but zygotes from oocytes with minimal cumulus developed less frequently (23%; P<0.001). In the third experiment, oocytes were classified as good or intermediate quality (omitting those with minimal cumulus), and presumptive zygotes of these groups were divided at 15 h after exposure to spermatozoa into those with dense even cytoplasm or thin and uneven cytoplasm. Cleaved zygotes from oocytes of good or intermediate quality developed to expanded blastocysts with similar frequency (49% vs 46%). Presumptive zygotes classified as having even cytoplasm, compared with those with uneven cytoplasm, had higher cleavage rates (87% vs 76%; P<0.001) and a higher frequency of development of cleaved zygotes to expanded blastocysts (53% vs 43%, P<0.001). The greatest gains in selecting oocytes and zygotes with high developmental potential were made by omitting oocytes with minimal cumulus and selecting presumptive zygotes with dense even cytoplasm.