Pre-implantation genetic diagnosis has become a powerful tool in human in vitro fertilization. Yet, for ethical reasons, controlled experiments assessing the effects of embryo biopsies on human embryos are difficult to perform. Therefore, a series of experiments was performed to examine the effects of blastomere biopsies on subsequent development of IVF-derived bovine embryos cultured under either atmospheric or low (5%) oxygen. Embryos were generated by IVF of in vitro-matured oocytes and cultured in CR1aa medium containing 5% bovine fetal serum. The first experiment was designed to assess the optimal time for blastomere removal. One blastomere was removed from each of a large number of embryos at either 48 or 72 h after IVF. Biopsy at 48 h resulted in 17.2% (10/58) of embryos proceeding to the blastocyst stage, which was significantly lower than when biopsies were performed at 72 h (37.5% (33/88), P < 0.05). In the second experiment, embryos were cultured under either atmospheric or 5% O₂ following blastomere removal. Biopsies at 72 h had no effect on rate of blastocyst formation with 38.5% (110/286) of controls and 33.7% (60/178) of biopsied embryos proceeding to the blastocyst stage. However, culture under 5% O₂ significantly increased the number of blastocysts from 29.9% (69/231) to 43.3% (101/233). This effect was significant in both biopsied and control embryos. In contrast, staining of blastocysts for cell numbers did not reveal any effects of biopsies or culture conditions. In the final experiment, biopsies were again performed at 72 h and embryos were cultured as previously. Blastocysts were collected and cultured individually for 48 h in 50-μL-medium droplets in their respective O₂ concentrations after which time the medium was assayed for concentration of interferon-tau (IFN-τ). Previous work has shown that IFN-τ secretion can be affected by genetic and environmental factors. Reduced O₂ concentration again significantly increased blastocyst formation from 24.9 to 36.9% (49/197 vs. 75/203, P < 0.05), while blastomere removal had no effect on development. IFN-τ secretion did not differ between biopsied and control blastocysts, although culture under atmospheric O₂ resulted in significantly increased IFN-τ concentration in medium droplets (12,690.1 ± 2715.6 vs. 4726.8 ± 2488.5 pM, P < 0.05).

This work was supported by a grant (HMK) from the National Institutes of Health (HD 36421).