TCR sequencing and cloning methods for repertoire analysis and isolation of tumor-reactive TCRs

T cell receptor (TCR) technologies, including repertoire analyses and T cell engineering, are increasingly important in the clinical management of cellular immunity in cancer, transplantation, and other immune dis-eases. However, sensitive and reliable methods for repertoire analyses and TCR cloning are still lacking. Here, we report on SEQTR, a high-throughput approach to analyze human and mouse repertoires that is more sensitive, reproducible, and accurate as compared with commonly used assays, and thus more reliably captures the complexity of blood and tumor TCR repertoires. We also present a TCR cloning strategy to spe-cifically amplify TCRs from T cell populations. Positioned downstream of single-cell or bulk TCR sequencing, it allows time-and cost-effective discovery, cloning, screening, and engineering of tumor-specific TCRs. Together, these methods will accelerate TCR repertoire analyses in discovery, translational, and clinical set-tings and permit fast TCR engineering for cellular therapies.

Automated summary

T cell receptor (TCR) technologies are essential in the clinical management of cellular immunity, but current methods for repertoire analyses and TCR cloning lack sensitivity and reliability. This study introduces SEQTR, a high-throughput approach that is more sensitive and accurate in analyzing human and mouse repertoires, enabling more reliable capture of the complexity of TCR repertoires. Additionally, a TCR cloning strategy is presented for efficient discovery, cloning, screening, and engineering of tumor-specific TCRs.