In the present study, we reinvestigated the effects of anti-Lyt-2 monoclonal antibodies (mAb) on the generation of H-2K/D-specific Lyt-2+ cytolytic T lymphocytes (CTL). A sensitive limiting dilution microculture system supplemented with a source of interleukin 2 (IL 2) was employed to provide a minimal estimation of the frequency of CTL precursors (CTL-P) in normal spleen or mixed leukocyte culture (MLC) populations. The addition of anti-Lyt-2 (but not anti-Thy-1) mAb at culture initiation caused an average 78% reduction in the number of CTL-P within spleen cell populations that were activated by allogeneic stimulation. Most CTL that were generated from Lyt-2+ CTL-P in the presence of anti-Lyt-2 mAb were not inhibited in cytolysis by such mAb, whereas the majority of CTL generated in the absence of anti-Lyt-2 were inhibitable. In contrast to normal spleen, the frequency of (operationally defined) CTL-P that had been activated for 5 days in primary MLC was unaffected by the presence of anti-Lyt-2 mAb. Collectively, these results demonstrate that anti-Lyt-2 mAb are capable of inhibiting the antigen-dependent activation of normal splenic precursors of Lyt-2+ CTL. The fact that IL 2 could not overcome the inhibition suggests that anti-Lyt-2 mAb may have affected the expression of IL 2 receptors as a direct consequence of inhibiting CTL-P antigen recognition.

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