A method for determining the sex of human skeletons was developed using molecular genetic techniques. The amelogenin gene, found on the X and Y chromosomes, was examined using the polymerase chain reaction (PCR) and a nonradioactive dot blot procedure. DNA was analyzed from 20 modern individuals of known sex and 20 skeletons from an archaeological site in central Illinois dating to A.D. 1300. An independent assessment of the sex of each skeleton was made according to standard osteological methods. The sex of 19 ancient and 20 modern individuals was accurately determined using this molecular genetic technique. Molecular sex determination will be especially useful for juvenile and fragmentary remains when it is difficult, or impossible, to establish an individual's sex from morphological features.