T cell coactivation is a dynamic process subject to integrin-dependent positive and negative regulation. Costimulation of human peripheral blood T cells by CD3 mAb OKT3 in conjunction with anti-alpha 4 has been shown to be down-regulated by the anti-beta 1.1 epitope-specific mAb 18D3. As expected, maximal costimulation induced by alpha 4-specific mAb L25 was inhibited (70%) by the addition of soluble mAb 18D3. Surprisingly, soluble mAb 18D3 inhibited maximal proliferation induced by the costimulatory alpha 4 beta 7-specific mAb ACT-1 by 40%, thus demonstrating that one integrin subfamily can regulate the activity of another. To determine whether mAb 18D3 could regulate more than alpha 4-associated integrin-mediated costimulation, non-alpha 4 integrins were tested. mAb 18D3 inhibited maximal proliferation induced by alpha 4-specific mAb 3D6, and an alpha 4-specific mAb 16. This clearly demonstrates that a variety of integrin costimulatory molecules (of the beta 1, beta 2, and beta 7 subfamilies) can be regulated negatively by mAb 18D3. To analyze the specificity of this negative regulation, other cell surface costimulatory molecules were tested for susceptibility to mAb 18D3. Although Abs specific for CD4, CD26, CD28, CD44, CD45RA, or CD45RO were sufficient to activate T cells when co-immobilized with anti-CD3 mAb, all were refractory to the inhibitory effects of mAb 18D3. Inhibition of T cell activation directly correlated with diminished IL-2 production. This suggests that mAb 18D3 selectively regulates integrin-dependent T cell activation by delivering a negative effect at some common point utilized by various integrin subfamilies.