Objective: To compare a single medium, with medium renewal on day 3, with a sequential media system for development of human embryos to the blastocyst stage and subsequent pregnancy outcome. Design: Prospectively randomized study using donor oocytes. Setting: Private infertility clinic. Patient(s): Oocytes were collected from 47 donors (mean ± SD age = 25.6 ± 3.5 years) from March to October 2006 and used for 79 patients (40.6 ± 5.4 years). The patients were prospectively randomized to have their embryos cultured in a single medium or in a sequential media system. Intervention(s): The donor oocytes were fertilized with fresh patient partner sperm and the resultant zygotes cultured in either a single medium or a sequential media system from day 1 until the blastocyst stage. After endometrial preparation the embryos were transferred to the patients on day 5 or day 6. Main Outcome Measure(s): In vitro embryo developmental characteristics and pregnancy and implantation rates. Result(s): In vitro development rates on days 3, 4, and 5 and implantation rates were significantly greater for embryos cultured in the single medium than for those cultured in the sequential media system. Conclusion(s): A single medium was as good as or better than a sequential media system for human embryo culture from the zygote to blastocyst stage. The idea that a sequential media system is required for optimal development of the human embryo to the blastocyst stage is highly questionable. © 2009 American Society for Reproductive Medicine.