A tumor-specific antigen (TSA) expressed on the chemically induced BALB/c Meth A sarcoma and one of 22 other BALB/c sarcomas tested, CMS13, was detected in in vitro cellular and humoral assays. The distribution pattern of the TSA defined in a complement-dependent microcytoxicity assay by cytotoxic antibodies present in CMS13 antisera was similar to that detected by a cytotoxic T-cell clone, designated CTLL-MA10B, in an 18-h cell-mediated cytotoxicity assay. The serologically defined TSA was shown to be expressed on gp96, a Mr 96,000 glycoprotein isolated from Meth A cytosol with immunoprotective activity in in vivo tumor rejection assays. Immunization of BALB/c mice with Meth A, CMS13, or preparations of gp96 isolated from these sarcomas induced tumor resistance in these mice to Meth A and CMS13 but not CMS5, an antigenically unrelated sarcoma. These results suggest that the shared TSA is expressed on gp96 and is functional in tumor rejection assays.