A method for the detection and direct typing of herpes simplex virus (HSV) by the polymerase chain reaction (PCR) technique has been developed. One common upstream primer and two type-specific downstream primers were prepared to amplify DNA from the HSV type 1 and type 2 DNA polymerase gene. Using these three primers simultaneously in the PCR reaction mixtures, both types of HSV DNA were amplified to produce products of different sizes. By direct gel analysis, the products of standard HSV type 1 and type 2 strains had the predictive sizes of 469 and 391 base pairs, respectively, and the difference in molecular mass enabled us to type the HSV strain. A total of 24 strains (type 1; 16 and type 2; 8 strains) were examined by PCR, and the results were consistent with those determined by immunofluorescence using type-specific monoclonal antibodies. No specific amplification was observed using other herpes virus or human genomic DNAs. The PCR method was then applied to clinical specimens. Of 15 samples obtained from oral lesions of children with herpetic gingivostomatitis, all (100%) were HSV positive by PCR, compared with 13 (86.7%) using standard cell culture methods. Three specimens from vulvar lesions of women with genital herpes were positive using both PCR and cell cultures. There was complete agreement in the typing of HSV strains using the PCR method or virus isolation. On the basis of these results, it is suggested that DNA amplification and typing by PCR is particularly useful for material from which virus isolation might be difficult.