analysis of the paired tcr α- and β-chains of single human t cells分析成对tcrα-β-chains单一人类t细胞.pdfVIP

Analysis of the Paired TCR a- and b-chains of Single Human T Cells 1 2,3 1 1 2,3 2 Song-Min Kim , Latika Bhonsle , Petra Besgen , Jens Nickel , Anna Backes , Kathrin Held , 1 2,3 1 Sigrid Vollmer , Klaus Dornmair , Joerg C. Prinz * 1 Department of Dermatology, Ludwig-Maximilian-University, Munich, Germany, 2 Institute of Clinical Neuroimmunology, Ludwig-Maximilian-University, Munich, Germany, 3 Max-Planck-Institute of Neurobiology, Martinsried, Germany Abstract Analysis of the paired i.e. matching TCR a- and b-chain rearrangements of single human T cells is required for a precise investigation of clonal diversity, tissue distribution and specificity of protective and pathologic T-cell mediated immune responses. Here we describe a multiplex RT-PCR based technology, which for the first time allows for an unbiased analysis of the complete sequences of both a- and b-chains of TCR from single T cells. We validated our technology by the analysis of the pathologic T-cell infiltrates from tissue lesions of two T-cell mediated autoimmune diseases, psoriasis vulgaris (PV) and multiple sclerosis (MS). In both disorders we could detect various T cell clones as defined by multiple T cells with identical a- and b-chain rearrangements distributed across the tissue lesions. In PV, single cell TCR analysis of lesional T cells identified clonal CD8+ T cell expansions that predominated in the epidermis of psoriatic plaques. An MS brain lesion contained two dominant CD8+ T-cell clones that extended over the white and grey matter and meninges. In both diseases several clonally expanded T cells c