Two major histocompatibility complex (MHC) class I-reactive T cell clones derived from H-2b mice, generated against the allogeneic Ld molecule, were found to recognize the H-2b class II mutant Abm12 molecule as well. In addition, these clones also recognize the class II A(s) molecule, and display a class II-dependent reactivity to staphylococcal enterotoxin B. Neither the class I nor the class II alloreactivities of the clones were found to be dependent on other MHC molecules. Both clones express CD4+CD8- phenotypes. The CD4 molecule appears to be involved in their class II reactivity, while little or no role for CD4 could be detected in the class I reactivity. This is the first report of a class I/class II cross-reactivity being mediated by CD4+ T cells. The structural basis for this cross-reactivity is discussed.