Mouse embryos at day 9 of development were cultured for 24 to 42 h in 50% fetal calf serum and 50% Waymouth's medium containing 0.5 micrograms/ml insulin supplemented with various amounts of progesterone and estradiol-17-beta. Unmodified medium contained approximately 0.2% of the normal maternal blood levels for that stage of pregnancy. The addition of 1 X 10(-7)M progesterone to the medium brought the level near that of the normal maternal circulating amount and appeared to be beneficial for in vitro development. After 24 h of cultivation there was a statistically significant increase in somite number, the number of embryos developing posterior limb buds, and protein accumulation over the control embryos. The addition of small amounts of estradiol-17 beta (1 X 10(-10)M) increased the protein accumulation of the embryo over that of progesterone alone and seems to enhance the beneficial effects of progesterone addition.