The pregnancy rates for the gamete intra-fallopian transfer (GIFT) and tubal embryo transfer (TET) methods have been shown to be high. The oviductal environment appears to be suitable for fertilization and subsequent cleavage. Many authors have reported that when co-cultured with human ampullary cells, the embryos can develop in vitro as well as in vivo. Thus, significantly more embryos reach the early blastocyst stage, and more successful implantation may be obtained. However, the characteristics of the ampullary cells may change after a series of sub-cultures. In this study, we compared the first and the seventh sub-culture ampullary cell co-cultured systems with a control group. There were significantly more embryos cleaved to blastocysts in these two co-cultured systems than in the control group, but no significant difference between these two co-cultured systems. Detoxification of the medium by co-cultured cells may play an important role in improving embryo quality.