Identification of antigen-specific TCR sequences using a strategy based on biological and statistical enrichment in unselected subjects

Recent advances in high-throughput T cell receptor (TCR) sequencing have allowed for new insights into the TCR repertoire. However, methods for capturing antigen-specific repertoires remain an area of development. Here, we describe a novel approach that utilizes both a biological and statistical enrichment to define putatively antigen-specific complementarity-determining region 3 (CDR3) repertoires in unrelated individuals. The biological enrichment entails fluorescence-activated cell sorting of in vitro antigen-activated memory CD4+ T cells followed by TCRβ sequencing. The resulting TCRβ sequences are then filtered by selecting those that are statistically enriched when compared to their frequency in the autologous resting T cell compartment. Applying this method to define putatively peanut protein-specific repertoires in 27 peanut-allergic individuals resulted in a library of 7345 unique CDR3 amino acid sequences that had similar characteristics to validated antigen-specific repertoires in terms of homology and diversity. In-depth analysis of these CDR3s revealed 36 public sequences that demonstrated high levels of convergent recombination. In a network analysis, the public CDR3s unveiled themselves as core sequences with more edges than their private counterparts. This method has the potential to be applied to a wide range of T cell mediated disorders, and to yield new biomarkers and biological insights.