Identification of antigen-specific TCR sequences based on biological and statistical enrichment in unselected individuals

Recent advances in high-throughput T cell receptor (TCR) sequencing have allowed for new insights into the human TCR repertoire. However, methods for capturing antigen-specific repertoires remain an area of development. Here, we describe a potentially novel approach that utilizes both a biological and statistical enrichment to define putatively antigen-specific complementarity-determining region 3 (CDR3) repertoires in unselected individuals. The biological enrichment entailed FACS of in vitro antigen-activated memory CD4(+) T cells, followed by TCR beta sequencing. The resulting TCR beta sequences were then filtered by selecting those that are statistically enriched when compared with 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 beta amino acid sequences that had similar characteristics to other validated antigen-specific repertoires in terms of homology and diversity. In-depth analysis of these CDR3 beta s revealed 36 public sequences that demonstrated high levels of convergent recombination. In a network analysis, the public CDR3 beta s were shown to be 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.

Automated summary

This study utilized a combination of biological and statistical approaches to define potentially antigen-specific CDR3 repertoires in unselected individuals through FACS of antigen-activated memory CD4(+) T cells and TCR beta sequencing. Application of this method to peanut-allergic individuals identified a library of unique CDR3 beta sequences with similarities to validated antigen-specific repertoires. Further analysis revealed public sequences with high levels of convergent recombination and network analysis showed these sequences to be core with more edges than private counterparts, indicating potential for use in T cell-mediated disorders and biomarker discovery.