To determine whether hormone analysis could be used for accurate determination of fetal sex, we measured testosterone (T) and follicle-stimulating hormone (FSH) in 130 amniotic fluid samples at midgestation. The mean unconjugated T in amniotic fluid of 73 patients carrying male fetuses was 202 pg. per milliliter (95 per cent confidence limits [CL]: 70 to 580) and all but three had levels higher than 90 pg. per milliliter. The mean amniotic fluid unconjugated T for 49 patients carrying female fetuses of 41 pg. per milliliter (95 per cent CL: 11 TO 125) was fivefold lower than that for the male fetuses and all but three patients carrying female fetuses had amniotic fluid T levels of 90 pg. per milliliter or lower. The mean amniotic fluid FSH of 0.7 ml. U. per milliliter (95 per cent CL: less than 0.5 to 3.4) for subjects with male fetuses was tenfold lower than that for patients with female fetuses. Amniotic fluid FSH levels less than 2.0 ml. U. per milliliter were found in 88 per cent of patients carrying male fetuses and in only one patient with a female fetus, and levels greater than 10 ml. U. per milliliter were found in those with female fetuses only. In eight patients (7 per cent of cases), neither amniotic fluid T nor FSH determinations were indicative of fetal sex. Measurement of unconjugated T and FSH in amniotic fluid may be an adjunct to other methods of determining fetal sex.