In an in vitro model, 20 condoms containing 0.9 mL of 6.6% (vol/vol) nonoxynol 9 and ten condoms without nonoxynol 9 were tested as physical and chemical barriers against human immunodeficiency virus (HIV). Each condom was mounted on a hollow dildo and placed in a glass cylinder. The HIV inoculum and HIV-free medium were placed on opposite sites of the condom. Intercourse was simulated by pumping the dildo up and down in the cylinder before and after deliberate rupture of the condom. Samples for HIV culture were taken from outside and inside the condom, before and after rupture. After rupture of nonoxynol 9-containing condoms, an outside nonoxynol 9 concentration of 0.25% was reached. No condom without nonoxynol 9 leaked HIV before rupture, but after rupture HIV could be detected in medium outside of seven of ten condoms tested. In none of 20 nonoxynol 9-containing condoms could HIV be detected in outside medium after rupture. Thus, undamaged condoms provide an effective physical barrier against HIV, and nonoxynol 9 may provide an effective chemical barrier as well.