Activation of naive CD4 T cells as measured by proliferation and 1L-2 secretion requires costimulation in addition to TCR ligation. To define the costimulatory signaling mediated by LFA-1 and its APC coreceptor ICAM-1 we have examined the effect of anti-LFA-1 mAb on both proliferation and cytokine differentiation using naïve T cells from DO 10 mice expressing the transgenic TCR specific for ovalbumin (OVA) peptide 323-339. Anti-LFA-1 inhibition of proliferation was determined at log intervals between 0.03 and 30 ng/ml OVA peptide using BALB/C (1-A") splenic adherent cells as APCs. The IC50 (|ig/ml) for anti-LFA-1 was less than 1 at peptide doses at or below that which gave maximal proliferation (3μg/ml OVA peptide) in the control but increased to approximately 9 at the highest antigen dose. Naïve T cells activated with either 3 or 30 |ig/ml OVA peptide in the presence of 10 ng/ml anti-LFA-1 were restimulated at various peptide doses at one week after the primary activation. Cytokine expression was assayed by in situ hybridization of mRNA and by ELISA. Blocking the LFA-1/ICAM-1 interaction in the primary activation induced a Th2-like phenotype in the secondary stimulation, characterized by increased production of IL-4 and IL-10 with concomitant inhibition of IFNy. These data suggest that LFA-1/ICAM-1 costimulatory signals are potent modulators of naïve T cell cytokine differentiation at antigen doses which maximize the proliferative response.