The use of three media to support growth and development in headfold Sprague-Dawley rat embryos cultured for 48 h, for possible use in a teratogen assay, was investigated using sheep serum (SM) and sheep serum mixed with rat serum (80% sheep:20% rat)(RSM). Rat serum (RM) was used as the control. The pCO2, p02, pH, bicarbonate concentration, glucose content and osmolarity of the media were determined at 0 h, 24 h and 48 h after the start of incubation. The embryos were assessed at 24 h and 48 h using yolk sac diameter, crown-rump length, somite number, protein content, stage of development and morphological scoring. Malformations were noted. The pCO2 and p02 levels in the media were similar to those present in the gas environment. The RM had a lower pH (P < 0.01), less bicarbonate (P < 0.01) and a higher osmolarity (P < 0.01) than SM and RSM, which had similar levels, although RSM had a higher osmolarity (P < 0.01) than SM. A drop in pH and bicarbonate level was noted over the culture period. Glucose levels dropped only in media containing embryos. For all parameters measured, the embryos cultured in RM were superior to those cultured in RSM and SM, with the embryos in SM being inferior to those of RSM in certain aspects. The embryos in RSM had more malformations per embryo than those in RM and SM. The RM produced fewer abnormal embryos (P < 0.01) than SM and RSM, which were similar. Most abnormalities detected involved the neural tube. In general, the embryos in RSM were not much improved on those in SM. In conclusion, sheep serum could not support acceptable normal embryonic growth and development for a teratogen- detect¬ing assay with the culture technique used in this study.