© Copyright 2015 Wolters Kluwer Health, Inc. All rights reserved. Background. Memory T cells (Tmem), particularly those resistant to costimulation blockade (CB), are a major barrier to transplant tolerance. The transcription factor Eomesodermin (Eomes) is critical for Tmem development and maintenance, but its expression by alloactivated T cells has not been examined in nonhuman primates. Methods. We evaluated Eomes and coinhibitory cytotoxic T lymphocyte antigen-4 (CTLA4) expression by alloactivated rhesusmonkey Tcells in the presence of CTLA4 immunoglobulin, both in vitro and in renal allograft recipients treated with CTLA4Ig, with or without regulatory dendritic cell (DCreg) infusion. Results. In normal monkeys, CD8+ T cells expressed significantly more Eomes than CD4+ T cells. By contrast, CD8+ T cells displayed minimal CTLA4. Among T cell subsets, central Tmem (Tcm) expressed the highest levels of Eomes. Notably, EomesloCTLA4hi cells displayed higher levels of CD25 and Foxp3 than EomeshiCTLA4lo CD8+ Tcells. After allostimulation, distinct proliferating EomesloCTLA4hi and EomeshiCTLA4lo CD8+ T cell populations were identified, with a high proportion of Tcm being EomesloCTLA4hi. CB with CTLA4Ig during allostimulation of CD8+ Tcells reduced CTLA4 but not Eomes expression, significantly reducing EomesloCTLA4hi cells. After transplantation with CB and rapamycin, donor-reactive EomesloCTLA4hi CD8+ Tcells were reduced. However, in monkeys also given DCreg, absolute numbers of these cells were elevated significantly. Conclusions. Low Eomes and high CTLA4 expression by donor-reactive CD8+ Tmem is associated with prolonged renal allograft survival induced by DCreg infusion in CTLA4Ig-Treated monkeys. Prolonged allograft survival associated with DCreg infusionmay be related to maintenance of donor-reactive EomesloCTLA4hi Tcm.