Development of a novel TCR avidity assay for human CD8+ T cells

T cells with high avidity for their specific ligand are believed to be most effective in clearing viral infections and inducing tumor regression. Thereby, an important part of the avidity seems to be hard wired in the structure of the T cell receptor (TCR). Existing methods do not allow for the exact measurement of this structural avidity on living cells. We therefore developed a novel assay, where the dissociation of monomeric MHC-molecules bound to surface expressed TCRs can be observed by real-time microscopy and can be quantified as koff-rate. Experiments with preclinical mouse models showed a clear correlation between functional avidity, koff-rate and protectivity of the T cells. This led to the assumption that the koff-rate might be an important parameter for determination of the quality of T cells used e.g. for adoptive immunotherapy. Hence, the assay was adapted to the measurement of human T cells. To achieve this, fundamental changes of the setups turned out to be necessary. Validation of the method with CMV-specific T cells showed that the assay allows for precise and reproducible determination of the koff-rate. High variability of the koff-rate assed for different CMV-positive donors indicates that this parameter might be important for the selection of high quality T cells for adoptive T cell therapy.