Figure 1. Analysis of lymphocytes from MBPβ transgenic mice cultured with the AC1-11 peptide.

Total lymphocytes from MBPβ-only transgenic mice were bulk cultured with the MBP-derived AC1-11 peptide. Several additional rounds of activation were done by culturing with T cell depleted B10.PL splenocytes loaded with the AC1-11 peptide. FACS was carried out seven days after the final activation. (A) Nearly all remaining cells were CD4⁺ and expressed the transgene-encoded Vβ8 TCRβ chain. Across several experiments, approximately 30% of the T cells were not Vα2 positive. (B) The Vα2⁺ and Vα2⁻ T cells were sorted and then stimulated by the addition of irradiated, T cell depleted B10.PL splenocytes and titrated amounts of the AC1-11 peptide. Cells were pulsed with 3[H]thymidine after 48 hours and harvested after a total of 72 hours. Approximately 5×10⁴ T cells and 1×10⁵ APCs were used per well. Assays were carried out in triplicate and averaged. Results shown are representative of three independent experiments. (C) Predicted amino acid sequence of the CDR3α region of the Vα6 (TRAV6-7/DV9*02) – Jα13 (TRAJ13*01) TCR cloned from the Vα2⁻ population. Gene nomeclature based on the IMTG Marie-Paule convention [46].