Induction of donor specific tolerance in composite tissue allografts (CTAs) is the ultimate goal in transplantation. Recently, we induced donor specific tolerance to rat hind limb allografts across MHC barrier under 35 days course of Cyclosporine A (CsA) and ab T-cell receptor (TCR) mAb therapy. In this report, we present the efficacy of shorter immunomodulating protocol and its effect on chimerism and tolerance induction. Thirty-six hind limb transplantations across MHC barrier, [Lewis-Brown-Norway (LBN, RT1^l+n) to Lewis (LEW, RT1^l)] were tested for potential to induce tolerance under 35, 21, 7 and 5 day protocols of combined CsA and ab TCR Mab therapy. There was no chronic immunosuppression once the protocol was stopped. Experimental groups included transplants under 4 different time/dose regimens and with monotherapy controls of CsA and TCR alone. Efficacy of immunomodulating therapy was measured by flow cytometry (FACS) levels of: CD3, CD4, CD8, CD90, NK and TCR. Clinical tolerance was tested by skin grafting from the donor (LBN), recipient (LEW) and third party (ACI, RT1^a). Donor specific chimerism was evaluated in the peripheral blood lymphocytes (PBL), and lymphoid organs by flow cytometry (FC). The expression of RT1l relative to the MHC class I antigen of the LBN donor’s CD4 and CD8 cells was assessed. Without immunosuppression limbs rejected at day 7. Monotherapy with CsA or ab-TCR, prolonged survival up to 21 and 13 days, respectively (p<0.05). All transplants (n=10) under 35 days of CsA/TCR protocol survived rejection-free over 700 days (p<0.0001). Limb allografts under short protocols of 5, 7, and 14 days are rejection-free and still under evaluation at 110, 125 and 150 days, respectively. Clinical tolerance was confirmed by acceptance of skin grafts from the CTA donors (LBN). This correlated with PBL chimerism of 12.6% of CD4+ and 6.8% of CD8+ chimeric RT1ⁿ⁺ T cell subpopulations. Evaluation of lymphoid organs confirmed higher levels of chimerism in splenic tissue (62.8% total, 11.8% being RT1ⁿ⁺/CD4+) and in para-aortical lymph nodes (14.7% total, 1.37% being RT1ⁿ⁺/CD4+), but not in thymic tissue. To the best of our knowledge, this is the first report documenting induction of tolerance across MHC barrier in CTA transplants submitted to short protocols of immunomodulatory therapy without a need for chronic immunosuppression. The mechanism of tolerance induction seems to be directly associated with the development of donor specific chimerism in peripheral blood and lymphoid organs. This protocol may have direct impact on management of clinical CTA’s.