The ex-vivo response of TCRα transgenic T cells show impaired IFNγ response and selection of a preferred TCRβ partner chain. (A) TCRα chain transgenic and littermate controls were immunized with PLP 56 to 70/CFA on Day 0 and PLP 56 to 70/IFA on Day 28. T cell proliferation is similar in the two groups. Data shown are mean values ± SE (TCRα transgenic (black bars), n = 4 mice at Day 10, 6 at Day 28 and 7 at Day 32; littermate controls (white bars), n = 8 mice at Day 10, 8 at Day 28 and 9 at Day 32). The data are representative of three independently performed experiments. (B) Immediately ex-vivo at Day 10, both groups make similar amounts of IFNγ, but at Day 28 no IFNγ response is seen in the TCRα transgenics (*P <0.05; Mann-Whitney U test) or subsequently after re-challenge at Day 32, (C) TCRα transgenics show an impaired IFNγ response compared to controls. (D) T cells from the TCRα chain transgenics were harvested at Day 32, cultured with peptide for 48 h and TCRβ chain CDR3 analysis of CD4+ CD69+ (n = 50 sequenced receptors from cDNA of bulk, sorted T cell lines) and CD4+ CD69− (n = 48 sequenced receptors from cDNA of bulk, sorted T cell lines) cells carried out. The TCRα chain transgenic impaired IFNγ phenotype correlates with clonal expansion of a dominant TCRβ chain (TRBV2, TRBJ2-5 CDR3 CASSQAGTGEDTQYF). (E) Spectratyping analysis of CD4+CD69+ TCRα chain transgenic and littermate control cells was carried out using V gene specific primers.