A murine monoclonal antibody (mAb; 7D6) that was mitogenic for T cells was derived from 129/Sv animals immunized with a T helper clone from C57BL/6 origin. Fluoresceinated 7D6 labeled T cells from most common mouse strains but not from 129/Sv and LP/J animals, and this labeling was inhibited by the anti-CD3 epsilon mAb 145-2C11. The mitogenicity of 7D6 for T cells had a similar strain specificity. The antibody immunoprecipitated the T cell receptor (TcR) complex from a T cell hybridoma. After dissociation of this immunoprecipitate with detergents, the CD3 gamma and epsilon chains were retained by the 7D6 antibody. Immunoprecipitation data were also obtained with COS cells transfected with the CD3 gamma, delta or epsilon chains alone, in pairs or together. They confirmed that 7D6 bound the CD3 gamma epsilon pair, suggesting that the antibody recognizes a conformational epitope formed by gamma epsilon pairing, whereas 145-2C11 bound both gamma epsilon and delta epsilon pairs. These results, therefore, add to current information about TcR structure and subunit stoichiometry. We have demonstrated that the 7D6 mAb specifically binds to a CD3 dimer comprised of gamma and epsilon chains. We thus provide additional evidence that indicates that two CD3 epsilon chains are found within the receptor, one linked to CD3 gamma and the other to CD3 delta.