Induction of antigen-specific T cell tolerance would aid treatment of diverse immunological disorders and help preventing allograft rejection and graft versus host disease. We established a method of inducing antigen-specific T cell tolerance in situ in diabetic humanized mice and non-human primates receiving porcine islet xenografts. Antigen-specific T cell tolerance was induced by administration of MD-3 antibody ligating a particular epitope on ICAM-1 (intercellular adhesion molecule 1). Antibody-mediated epitope-based ligation of ICAM-1 on dendritic cells (DCs) led to the arrest of DCs in a semimature stage in vitro and in vivo. Ablation of DCs from mice completely abrogated anti–ICAM-1–induced antigen-specific T cell tolerance. T cell responses to unrelated antigens remained unaffected. When MD-3 antibody was applied with anti-CD154 antibody and rapamycin in diabetic Rhesus macaques, this combined therapy allowed the survival of porcine islet xenograft approximately for 1 year. Through meticulous analysis of T cell-dependent and –independent immune response in non-human primates that received porcine islets, we found indispensable role of MD-3 in induction of T cell tolerance, whereas anti-CD154 antibody therapy was not able to sufficiently suppress T cell response in xenograft setting. Rather, anti-CD154 antibody showed excellent effect in control of T-independent humoral response, when used in combination with rapamycin that effectively suppresses NK cell activation. This MD-3-based therapy did not induce any intractable adverse effects. Taken together, we expect that this approach might enable the achievement of transplantation tolerance and thereby successful islet xenotransplantation in clinical context.