The usefulness of the polymerase chain reaction (PCR) method for diagnosing adenovirus infections was investigated. Several primers, including primers specific for the hexon-coding region and for enteric adenovirus types 40 and 41, were evaluated. The PCR method was validated against cell culturing in routine diagnostic work and against restriction enzyme analysis of viral DNA. Sixty diagnostic specimens were selected for evaluation by the PCR method. Twenty of the 60 specimens were found positive on the basis of cytopathic effects and latex agglutination (Adenolex [Orion Diagnostica, Helsinki, Finland]), and 16 were identified and typed as adenoviruses by polyacrylamide gel electrophoresis. PCR was performed on all 60 specimens in parallel directly on diluted stool samples and on viral DNA extracted from cells inoculated with the same stool samples. When the general hexon primers were used 51 of the 60 specimens from infected cell cultures were found positive by PCR, whereas only 13 specimens were found positive when PCR was performed directly on stool samples. With the use of selective primers for enteric adenoviruses 16 of the 60 cell cultures were found to exhibit amplification products by PCR, whereas 4 were detected in stool samples. None of the 60 specimens were found positive by PCR when an adenovirus type 40-specific primer pair was used. PCR was found to be a fast, sensitive, and reliable method for the detection of adenoviruses in diarrheal disease, provided the amplifications were performed directly on diluted stool samples.