Figure 2.

Enumeration of InsB:DQ8 tetramer⁺ cells from HLA-DQ8⁺ patients with T1D and individuals without diabetes. A: Representative flow cytometry plots showing dual InsB:DQ8 tetramer⁺ stained cells concatenated from six patients with T1D patients (left) and four concatenated DQ8⁺ individuals without diabetes (right). Cells were gated on size, single cells, live, CD20⁻, CD33⁻, CD123⁻, CD8α⁻, CD3ε⁺, CD4⁺. B: Frequency of insulin-tetramer⁺ cells/1 million CD4⁺ T cells from all patients with T1D (n = 37) and control subjects without diabetes (n = 26; biological replicates). C: Correlation of the frequency of tetramer⁺ cells with disease duration for all patients with T1D. D: Comparison of the frequency of tetramer⁺ cells in individuals with T1D duration shorter and longer than 5 years. E: Flow cytometry plots showing dual InsB:DQ8 (top) and HA:DR4 (bottom) tetramer staining within peripheral blood CD4⁺ T cells from five DR4⁺/DQ8⁺ subjects with new-onset after 10 days of in vitro stimulation with modified InsB_10–23 or HA_306–318 (data are from two independent experiments). Bars represent the mean. The Mann-Whitney U test for nonparametric data (B and D) or linear regression (C) was used to determine significance. PE, R-phycoerythrin.