Ivf plus medium

More than half of the patients received ovarian stimulation with GnRH analogues, and the rest were treated without GnRH analogues for controlled ovarian hyperstimulation. Insemination was performed in IVF plus medium (Vitrolife, Sweden) using conventional IVF. Approximately 16–18 h after insemination, oocytes with 1 or 2 visible pronuclei were further cultured in G1 plus medium (Vitrolife, Sweden). On day 3 of culture, embryos were scored as grade I, II, III or VI based on the number, size, and shape of blastomeres and their degree of fragmentation, as previously described^{32 (link)}. Grade VI embryos were discarded. D3 embryos were subjected to transfer, frozen by vitrification technology or expanded in G2-plus culture medium (Vitrolife, Sweden). Blastocysts from day 5 or 6 were scored using the system of Gardner. The usable blastocysts were frozen by vitrification technology or transferred.

After oocytes retrieval, insemination was performed by the conventional IVF or ICSI method in IVF-plus medium (Vitrolife, Sweden). The observation of pronuclei, embryo culture and score were performed as we described previously^{24 (link)}. Briefly, 18 h after insemination, zygotic embryos (day 1) with 2 pronuclei were considered normal fertilization and were placed into G1 plus medium (Vitrolife, Sweden) for further culture. On day 3, the morphology of the embryos was evaluated and scored according to a previously described method^{24 (link)}. Grade I and II embryos were taken as top embryos, while Grade III embryos were taken as non-top embryos. These embryos were frozen by vitrification, transferred into the uterine cavity, or further cultured to the blastocyst stage. For further culture, day 3 embryos were placed into G2-plus medium (Vitrolife, Sweden), and on day 5 or 6, the embryos were morphologically evaluated and scored. The usable blastocysts were frozen by vitrification or transferred into the uterine cavity. Morphological evaluation of embryos in our reproductive centre was performed by Dr Yufeng Wang, who has more than ten years of work experience as an embryologist.