High avidity TCR/pMHC binding results in the deletion of thymocytes by negative selection. However, for natural regulatory T-cell (TR) precursors, high avidity TCR/pMHC interaction leads to differentiation and escape from negative selection. The process of diverting thymocytes into alternative pathways of development is incompletely understood. In this study, we consider the role that pMHC density may have in this process. Using two TCR transgenic mice against differentially presented Hen egg white lysozyme (HEL) epitopes, we evaluated negative selection and Foxp3+CD4SP T cell development in response to graded amounts of intravenously injected HEL. In both mice, thymocyte sensitivity to negative selection and TR induction depended on antigen dose and reflected the level of presentation of their cognate epitopes. The absolute increase in TR occurred at doses of HEL lower than the dose required for complete negative selection of conventional CD4SP. TR induction was inefficient in the absence of TCRα gene rearrangement and was not a result of TR subset expansion. Our data shows that pMHC density provides an important signal in the routing of TR precursors away from negative selection towards development as regulatory T-cells.