We investigated the function of the CD8 moiety in antigen-specific and alternative activation of HLA class I- and HLA class II-allospecific CD8+ cytotoxic T lymphocyte (CTL) clones. Monoclonal antibodies (mAb) directed against the CD8 structure were only found to inhibit antigen-specific cytotoxicity of class I-allospecific CD8+ CTL clones and not of a class II-allospecific CD8+ CTL clone. However, cytotoxicity induced by CD3 mAb (used at suboptimal concentrations) or CD2 mAb in both types of CTL clone was blocked by CD8 mAb. The class II-allospecific CD8+ CTL clone was uniformly more difficult to inhibit than the class I-allospecific CD8+ CTL clones and, moreover, also easier to induce to exert nonspecific cytotoxicity by CD2 mAb and CD3 mAb. The absence of CD8 mAb blocking of antigen-specific cytotoxicity of the class II-specific CD8+ CTL clone is, therefore, assumed to result from too strong a triggering signal to be overcome by the down-regulatory signal of the CD8 antigen. These combined findings suggest a down-regulatory function of CD8 not only in T cell receptor (TcR)/CD3 activation, but also in TcR/CD3-controlled alternative activation routes such as the CD2 activation pathway.