Activated T cells and their naive precursors display different functional avidities for peptide/MHC, but are thought to have identical antigenic repertoires. We show that, following activation with a cognate mimotope (NRP), diabetogenic CD8+ T cells expressing a single TCR (8.3) respond vigorously to numerous peptide analogs of NRP that were unable to elicit any responses from naive 8.3-CD8+ T cells, even at high concentrations. The NRP-reactive, in vivo activated CD8+ cells arising in pancreatic islets of nonobese diabetic mice are similarly promiscuous for peptide/MHC, and paradoxically this promiscuity expands as the aviditiy of the T cell population for NRP/MHC increases with age. Thus, activation and avidity maturation of T lymphocyte populations can lead to dramatic expansions in the range of peptides that elicit functional T cell responses.