17 beta-Estradiol (E2) stimulated the zona-free hamster ova penetration by human spermatozoa from both fertile and subfertile men after 5 hours of incubation in vitro. For spermatozoa samples from fertile men, the mean penetration rates increased significantly (P less than 0.05), from 35.6% +/- 3.5% (control) to 60.1% +/- 5.8% (E2 = 50 ng/ ml) and 50.5% +/- 2.6% (E2 = 100 ng/ml), respectively. E2 also increased the mean penetration rates of spermatozoa samples from subfertile men significantly (P less than 0.05), from 10.2% +/- 1.8% (control) to 30.5% +/- 7.2% (E2 = 50 ng/ml) and 33.1% +/- 6.6% (E2 = 100 ng/ml), respectively. These findings indicate that E2 can affect the fertilizing ability of human spermatozoa in vitro.