New monoclonal anti-MyoD1 and anti-myogenin antibodies were evaluated immunohistochemically to determine whether they are useful in discriminating rhabdomyosarcoma (RMS) from other soft tissue tumors in routinely processed sections. Neither MyoD1 nor myogenin was expressed in normal, mature striated muscle. In RMS, nuclear expression of MyoD1 and myogenin was found in 82 and 80% of non-overlapping cases, respectively. MyoD1 was generally expressed in small, primitive tumor cells, and larger cells exhibiting morphological evidence of skeletal muscle differentiation failed to express positive nuclear immunostaining. Positive nuclear staining for myogenin was stronger than that for MyoD1 in cases with abundant differentiated tumor cells, but was less prominent in cases in which small, primitive tumor cells predominated. No leiomyosarcomas, Ewing's sarcomas/peripheral primitive neuroectodermal tumors or other soft tissue tumors exhibited nuclear expression of MyoD1 or myogenin. In conclusion, both anti-MyoD1 and anti-myogenin antibodies are useful for diagnosing RMS and for discriminating RMS from other soft tissue tumors.