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We have investigated the binding potential of staphylococcal enterotoxin B (SEB) to murine T cells using the induction of early activation events in Th1 and Th2 T-cell clones in the absence of antigen-presenting cells (APC) as indicators of direct interactions between SEB and the T-cell receptor (TcR). We consistently found that concanavalin A (Con A) induced rises in intracellular free calcium as well as inositol phosphate accumulation in APC-free T-cell clones. However, SEB uniformly failed to induce either calcium fluxes or inositol phosphate turnover in Th1 and Th2 T-cell clones in the absence of APC. In addition, we have used proliferation assays to show that (i) T-cell clones prepulsed with SEB did not respond when APC were added, (ii) APC-independent T-cell clones responded to soluble anti-TcR antibodies but not to SEB in the absence of APC, and (iii) SEB coupled to Sepharose beads did not stimulate T-cell clones in the absence of APC. Taken together our results argue against SEB binding to the TcR without the participation of MHC class II molecules. [ABSTRACT FROM AUTHOR]