T-cell Ig and mucin domain-containing protein (Tim)-2 is associated with Th2-dependent immune responses. Here, we investigated whether administration of anti-Tim-2 Abs affects the development of murine experimental allergic conjunctivitis (EC), a Th2-mediated disease. While treatment with anti-Tim-2 Abs in vivo during the induction phase did not affect the severity of EC, treatment during the effector phase augmented EC. The in vitro stimulation of ragweed (RW)-primed splenocytes with RW in the presence of anti-Tim-2 Abs induced significantly more IL-5 and IL-13 production but significantly less IFN-gamma production compared with the control splenocytes treated with normal rat IgG. In addition, the transfer of the anti-Tim-2 Ab-treated splenocytes induced significantly more severe EC than the control splenocytes. Thus, Tim-2 negatively regulates the Th2 differentiation of Ag-primed T-cells and the development of EC during the effector phase.