IMPROVED YIELDS OF BOVINE BLASTOCYSTS FROM IN-VITRO PRODUCED OOCYTES .2. MEDIA AND COCULTURE CELLS

Three experiments were conducted with 9,472 oocytes, aspirated from ov aries of slaughtered cows (7,318 cleaved zygotes), to compare some med ia and co-culture cells for in vitro maturation of oocytes and culture of embryos. The endpoint was development to expanded blastocysts at 9 .0 d after exposure of matured oocytes to sperm. In the first experime nt, oocytes were matured in TCM-199 and co-cultured in TCM-199 or Mene zo's B2 (B2), with granulosa or buffalo rat liver (BRL) cells. The typ e of co-culture cell had no effect on the proportion of oocytes or cle aved zygotes that developed to expanded blastocysts, but 31% of cleave d zygotes developed to expanded blastocysts in B2 compared with 21% in TCM-199 (P<0.001), and embryos developed faster in B2. In the second experiment, oocytes were matured in TCM-199, Ham's F-10 (F-10), or B2, then all were co-cultured in B2 with BRL cells. The proportion of cle aved zygotes that developed to expanded blastocysts was 28%, 40%, and 8% after maturation in TCM-199, F-10, or B2, respectively (P<0.001). I n the third experiment, presumptive zygotes were vortexed to remove cu mulus at 15 hr after exposure to sperm and classified as having dense even cytoplasm or thin and/or uneven cytoplasm. Then zygotes were co-c ultured with BRL or bovine oviduct epithelial (BOE) cells. There was l ittle difference in the proportion of cleaved zygotes that developed t o expanded blastocysts after coculture with BRL (51%) or BOE cells (54 %), but embryos developed somewhat faster with BOE than with BRL cells . Presumptive zygotes with even cytoplasm, compared to presumptive zyg otes with uneven cytoplasm, had higher cleavage rates (85% vs 76%, P<0 .001) and higher rates of development of cleaved zygotes to expanded b lastocysts (59% vs 44%, P<0.001). In these experiments, the best combi nation for growing embryos in vitro was F-10 maturation medium, B2 emb ryo culture medium, and BOE cells for co-culture.