Epidermal growth factor may play an important part in postnatal gastrointestinal development. However, little is known of its role prenatally. The aim of this study was to detect epidermal growth factor in amniotic fluid and to study its uptake and transfer across the epithelium of fetal rat small intestine. Anesthetized 20-day gestation rats underwent caesarian section. Three fetuses were exteriorized, their abdomens were opened, and ligated loops of proximal and distal small intestine were infused with 100 μg epidermal growth factor. Infused segments were removed 30 min later and processed for electron microscopy, and tissue was embedded in LR gold resin. Sections were treated with rabbit anti-rat epidermal growth factor antibody, followed by 5 nm gold-labeled goat anti-rabbit immunoglobulin before staining. Epidermal growth factor was measured in amniotic fluid by radioimmunoassay. In the proximal and distal small intestine epidermal growth factor was found membrane-associated along the luminal surface of microvilli, within apical invaginations and endosomal compartments, free from the membrane in multivesicular bodies, within large clear vesicles, basal vesicles and in association with the basolateral membrane and beyond. Epidermal growth factor was found in amniotic fluid at a concentration of 0.38 ± 0.07 (SD) ng/ml. This study shows that epidermal growth factor is present in amniotic fluid and is transported across the epithelium of fetal rats by an endocytotic process in both the upper and lower small intestine. It is likely that amniotic fluid epidermal growth factor plays a part in intestinal mucosal development, and may be active systemically after transepithelial passage in utero. © 1990.