Preliminary clinical experience with human blastocyst development in vitrowithout co-culture

This preliminary analysis was designed to quantify blastocyst development o f supernumerary embryos without the use of feeder cells, conditioned medium or whole serum. Embryos derived from in-vitro fertilization (IVF) that wer e not transferred or cryopreserved were included in this study. Ova were ha rvested for IVF after a standard ovarian stimulation with gonadotrophin-rel easing hormone agonist/human menopausal gonadotrophin (GnRHa/HMG) or follic le-stimulating hormone (FSH). Ova were collected and cultured in 150 mu l d roplets of PI medium under mineral oil, in groups at 37 degrees C under 5% CO2, 5% O-2, 90% N-2 (group A) or under 5% CO2 in air (group B) environment . Embryo transfer was performed 72 h post-harvest. Viable embryos not trans ferred or cryopreserved were placed in blastocyst medium and cultured for a n additional 48 h in 5% CO2 in air. Embryos that exhibited an expanded blas tocoelic cavity and well-defined inner cell mass at 120 h were counted. Of 838 supernumerary embryos cultured, 448 (53.5%) reached the expanded blasto cyst stage by 120 h of culture. Patients were given the option of cryoprese rvation at that time. The embryos were cryopreserved using a standard proto col with serial addition of glycerol, Embryos reaching the blastocyst stage after more than 120 h of culture were not included, There was no differenc e in the proportions of blastocyst development between group A, 217/410 (53 .5%) and group B, 231/428 (54%), To date, 16 patients have each had up to t hree thawed blastocysts transferred, out of whom seven became pregnant. Thi s report demonstrates that a simple system of sequential culture generated acceptable, viable blastocyst development (54%) with supernumerary embryos, without the use of feeder cells, conditioned medium or whole serum. Recogn izing the differential metabolic requirements of early and late cleavage st age embryos has enabled the application of a glucose/phosphate-free simple culture medium (P1) for up to 72 h of culture and a complex, glucose-contai ning medium (blastocyst medium) for subsequent blastocyst development.