Identifying and Tracking Low-Frequency Virus-Specific TCR Clonotypes Using High-Throughput Sequencing

Tracking antigen-specific T cell responses over time within individuals is difficult because of lack of knowledge of antigen-specific TCR sequences, limitations in sample size, and assay sensitivities. We hypothesized that analyses of high-throughput sequencing of TCR clonotypes could provide functional readouts of individuals' immunological histories. Using high-throughput TCR sequencing, we develop a database of TCRβ sequences from large cohorts of mice before (naive) and after smallpox vaccination. We computationally identify 315 vaccine-associated TCR sequences (VATS) that are used to train a diagnostic classifier that distinguishes naive from vaccinated samples in mice up to 9 months post-vaccination with >99% accuracy. We determine that the VATS library contains virus-responsive TCRs by in vitro expansion assays and virus-specific tetramer sorting. These data outline a platform for advancing our capabilities to identify pathogen-specific TCR sequences, which can be used to identify and quantitate low-frequency pathogen-specific TCR sequences in circulation over time with exceptional sensitivity.