These studies show that the probe EFD52 is suitable for forensic casework. No unexpected DNA banding patterns were obtained from controlled studies that examined various tissues, sample consistency over many gels, mixtures of body fluids, various substrates, various contaminants, and nonhuman DNA sources. Of all the animal DNAs tested, only one higher primate yielded a single band to EFD52 hybridization. The sensitivity of EFD52 was shown to be comparable to that of other forensic probes. Population frequency distribution tables were prepared from over 4,000 alleles, and two-locus studies were conducted on nine forensically useful probes. Black, white, Hispanic, and Lumbee Indian populations were found to be in Hardy-Weinberg and linkage equilibrium. Comparisons between victim blood standards and epithelial fractions of mixed stains from sexual assault cases were used to demonstrate the robustness of the EFD52 probe in forensic casework. 12 tables, 4 figures, and 32 references