CD45 is a family of transmembrane protein tyrosine phosphatases exclusively expressed by hematopoietic cells and critically involved in the regulation of T cell activation signals. We now demonstrate that three 100-μg doses of anti-CD45RB mAb MB23G2 can induce long-term engraftment of islets into major histcompatibility complex-disparate chemically diabetic mice. Long-term graft survivors (>120 days) were tolerant to new islet allografts from the original donor strain. MB23G2 induced a temporary decrease in number circulating leukocytes but had no effect on leukocyte number in other lymphoid compartments. Histologic examination of allografts from treated and untreated recipients revealed a similar peri-islet infiltration on day 6. Eleven days after transplant, the peri-islet infiltrate in treated animals persisted, but in marked contrast to untreated control animals, there was no insulitis and islet integrity was preserved. The peri-islet infiltrate from treated animals showed a mild increase in CD4 cells, a decrease in CD8 cells, and decreased intensity of CD45RB expression. Treatment of naive animals with anti-CD45RB (MB23G2) resulted in a shift in CD45 isoform expression on T cells with a loss of higher molecular weight isoforms and increased expression of lower molecular weight (CD45R0) isoform. This shift in CD45 isoform expression from CD45RB^Hi to CD45RB^Lo was associated with an increase in the intragraft expression of transcripts for interleukin (IL) 4 and IL-10, consistent with the expected activity of this distinct immunoregulatory T cell subset. Antibody-mediated targeting of CD45 may induce tolerance through novel mechanisms and have direct applicability to clinical transplantation in humans.