Human fetal adrenal tissue (from fetuses 9–19 weeks old) was studied for Δ5-3β-hydroxysteroid dehydrogenase activity by 3 methods: (a) conversion of dehydroepiandrosterone to Δ4-3-ketosteroids during in vitro incubation of tissue homogenates; (b) enzymatic assay based on the rate of androst-4-ene-3,17-dione formation from dehydroepiandrosterone; (c) enzymatic assay based on histochemical methods. Δ5-3β- Hydroxysteroid dehydrogenase activity was demonstrated by each of the 3 methods. The activity varied markedly among individual adrenals without any correlation with fetal age within the period studied. The absolute levels were of the same order of magnitude as those present in human adult adrenal tissue, with activity present as early as the 9th week of gestation. Δ5-3β-Hydroxysteroid dehydrogenase activity was also found in testis tissue removed from a 12-week-old fetus.