Distinct epitopes are recognized by cytolytic T lymphocyte clones on the same class I molecule: direct demonstration using DNA‐transfected targets and longterm cytolytic T cell clones

By producing long‐term, stable, cytolytic T lymphocyte clones and utilizing targets expressing only a single gene product derived from the stimulator mouse strain, we have been able to directly demonstrate that T cells recognize distinct epitopes expressed on a single H‐2 molecule. These multiple determinants are distinguishable by inhibition patterns with monoclonal antibodies (mAb). When two T cell clones, P‐2.14 and P‐2.17, are tested on an L cell transfected with the D^p gene (λ12a), the T cells kill the transfected targets as well as blasts derived from D^p mouse strains. mAb 7‐16.10 inhibits recognition and killing of D^p targets by both P‐2.14 and P‐2.17. This mAb 7‐16.10, however, competees with H‐2.m22. Interestingly mAb 11‐20.3 which also recognizes the H‐2.m22 specificity inhibits clone P‐2.14 but not P‐2.17. The mAb 7‐16.10, however, competes with 11‐20.3 for binding to the surface of L cells expressing the D^p gene. Thus the two T cells must recognize an overlapping specificity. Other mAb which bind to the H‐2D^p molecule are unable to inhibit either of these two cytolytic T cell clones. Paradoxically, any of the mAb when tested individually are sufficient to inhibit the polyclonal response derived from in vitro mixed lymphocyte culture. Therefore, by using targets expressing only a single H‐2 molecule derived by DNA‐mediated gene transfer and cytolytic T cell clones we have been able to directly demonstrate the presence of multiple epitopes on a single molecule and define their inhibition with mAb directed to that same molecule.