Abstract Suppression of patients’ T-cells is a key event in CLL pathogenesis and was demonstrated to be mediated by direct cell-cell contact of malignant CLL cells with T-cells. CD200 plays a critical role in regulating the immune system and has been shown to be up-regulated on the surface of different tumors including CLL. In this study we addressed the effects of CD200 over-expression on CLL cells on autologous T cells in a mixed lymphocyte reaction system. We used native and CD40 ligand (CD40L)- stimulated CLL cells as antigen-presenting cells (APCs) to expand autologous T cells of 14 patients. T-cell proliferation was analyzed over 3 weeks of in vitro culture. A functional anti- CD200 antibody (1B9) was added to reveal CD200-mediated immunosuppression in the autologous system. Expansion of patient T-cells was assessed by flow cytometry including intracellular staining of FOXP3. Specificity towards CLL-specific antigens was monitored applying fibromodulin derived peptides for detection of specific T-cells by ELISPOT analysis. T-cell proliferation over 3 weeks of in vitro culture was significantly enhanced compared to control cells when using CD40L-stimulated APCs and an anti-CD200 antibody (p=0.0004). CD200 blockade was further shown to stimulate antigen-specific T-cell responses towards the F2 and F4 peptides of the CLL-associated antigen fibromodulin (p=0.04). Finally, the number of CD4+/CD25high/FOXP3+ T cells (Treg) was significantly decreased in CD200 treated mixed lymphocyte reaction (p=0.04). In summary, CD200 blockade may provide therapeutic benefits in CLL by enhancing T-cell expansion, augmenting an antigen-specific T cell response with suppression of regulatory T cells. CD200 seems to be an important immunosuppressive molecule in CLL: by CD200 blockade immune suppression can be overcome by altering tolerance to tumor antigens and deregulation of regulatory T cells. This combination of an immune induction paralleled by a disruption of immunosuppressive factors makes anti-CD200 mAb a powerful tool for future treatment of CLL, possibly in combination with other B cell cytotoxic or immunostimulatory approaches.