ICAM-3 (CD50) is a panhematopoietic, constitutively expressed adhesion molecule which binds to its ligand LFA-1 by extracellular domain 1. ICAM-3 mediates the initial adhesion of T lymphocytes to LFA-1 bearing monocytes and B cells. Prior reports have demonstrated contradictory T cell activation results; crosslinking surface ICAM-3 and CD3 with mab have been shown to cause both stimulation and inhibition. We show that co-aggregation of ICAM-3 and CD3 in freshly isolated human T cells with immobilized mab significantly increased phosphatidylinositol (PI) hydrolysis (p<0.001), significantly increased cell size consistent with blastogenesis, significantly increased surface expression of CD69 and CD25 and significantly increased cellular metabolism (p<0.001) when compared with CD3/ICAM-3 negative isotype control mab. In addition, simultaneous CD3 aggregation and stimulation with mab to ICAM-3's extracellular domain 2 significantly enhance PI hydrolysis (p<0.001), indicating both extracellular domains are functional. We conclude ICAM-3 is an important T cell co-stimulus.