The CD28/B7 system provides costimulatory signals necessary for optimal T cell activation. We have examined the effects of blocking B7. 1 and/or B7. 2 in an in vitro system using TCR transgenic T cells specific for myelin basic protein. Activation of naive T cells was found to be B7. 2 dependent and not dependent on the presence of B7. 1 molecules. However, increasing the strength of signal through the TCR using peptide analogues with higher affinity for MHC compensated for blockade of B7. 2 molecules, suggesting that signal 1 alone can be sufficient for the activation of naive T cells. The role of B7 molecules in the differentiation of T cells was further investigated by restimulating T cells with fresh APC and peptide in B7-sufficient conditions. A down-regulation of IL-2 and IFN-γ production by T cells primed in the presence of anti-B7. 2 mAb was partially overcome when high affinity peptide analogues were used to restimulate T cells. In contrast, a significant down-regulation of the differentiation of cells producing Th-2 cytokines was observed in the presence of anti-B7 Abs. Differentiation of IL-4-secreting cells was influenced by both B7. 1 and B7. 2, while IL-5 secretion was totally dependent on B7. 2. These results suggest that B7-mediated costimulation is essential for the development of Th-2-associated cytokines, the absence of which cannot be overcome by increasing the strength of the signal through the TCR.