Abstract Tregs are professional suppressors of the immune response, yet their mechanism of action in vivo remains unclear. We compared the morphology and stoichiometry of the interactions of activated 5CC7 T cells and 5CC7 iTregs with MCC88–103 pulsed splenic DCs by electron and confocal microscopy. Image analyses revealed that Tregs interacting with DC displayed a distinct morphology with finger like membrane projections at the DC binding site and uropods at the rear end within three hours of in vitro culture. In contrast, activated T cells maintained their round morphology. Tregs occupied a greater extent of the DC surface than activated T cells consistent with a higher binding avidity. Dynamic intravital two-photon microscopy of adoptively transferred OTII iTregs and activated OTII cells demonstrated that Treg displayed a greater volume and duration of contact with OVA323–339 pulsed DC compared to that of activated OTII cells. Subsequent to their high avidity interactions with DC, 5CC7 iTregs captured MCC88–103-I-Ek complexes from DC surface reducing the amount of MCC presented on DC. Reduced antigen presentation was not due to global suppression of the DC’s ability to present antigen as 5CC7 and 3A9 iTregs only captured their cognate peptide-MHCII from MCC/HEL double pulsed DC, reducing the DC presentation in an antigen specific manner. When double pulsed DCs were cultured with iTregs specific for one peptide, separated from the iTregs, the DC failed to prime T cells specific for the antigen seen by the iTreg, but primed T cells specific for the second antigen. Altogether, we propose antigen specific depletion of peptide-MHCII complexes as a new mechanism for Treg-mediated suppression.