This study determined the prevalence of human papillomavirus 16/18 DNA in deparaffinized oral carcinoma specimens on slides with the use of the different sensitivities of in situ hybridization and a technique that combines polymerase chain reaction and in situ hybridization. Human papillomavirus DNA was not detected in the 30 biopsy specimens analyzed by in situ hybridization alone using biotinylated DNA probes specific for human papillomavirus 16/18. Twenty of 30 specimens (66.7%) were found to have human papillomavirus DNA (p < 0.001) with the use of the polymerase chain reaction-in situ hybridization technique. Human papillomavirus 16 was detected in 18 of 26 specimens (69.2%), and 7 of 25 carcinomas (28%) were found to contain human papillomavirus 18. Dual infections were present in 5 of 21 (23.8%) specimens. Human papillomavirus DNA was more prevalent in men (75%) than women (57.1%). However, there was no difference in the mean age of patients with oral carcinoma (men, 67.8 years; women, 67.5 years) who had human papillomavirus and those who did not (67.2 years). Carcinomas associated with dual infections occurred at a lower mean age (59.4 years) than those associated with a single human papillomavirus type (p < 0.005). We conclude that the polymerase chain reaction-in situ hybridization technique enhances our ability to demonstrate human papillomavirus types highly associated with oral squamous cell carcinoma.