The DNA probe and polymerase chain reaction (PCR) technique for detection and identification of mycobacteria were compared with the conventional smear and culture method. The results of identification by DNA probe agreed well with those of the biochemical method. Moreover, six percent of Mycobacterium avium complex (MAC) were revealed to be mycobacteria other than MAC by DNA probe. The nested PCR for detection of gene coding protein antigen b of Mycobacterium tuberculosis complex showed excellent specificity and sensitivity. Then we applied this technique to rapid detection of M. tuberculosis in 222 clinical samples. The agreement between nested PCR and the biochemical method was excellent, and 17 cases were diagnosed by only nested PCR in spite of negative results by smear and culture. These cases were unlikely to have yielded false positive results since their clinical features were compatible to tuberculosis. From these data, it was considered that the DNA probe and PCR technique were extremely useful strategies and would contribute to rapid diagnosis of mycobacterial infectious diseases.