Defining T cell receptor repertoires using nanovial-based binding and functional screening
Doyeon Koo, Zhiyuan Mao, Robert Dimatteo, Miyako Noguchi, Natalie Tsubamoto, Jami McLaughlin, Wendy Tran, Sohyung Lee, Donghui Cheng, Joseph de Rutte, Giselle Burton Sojo, Owen N. Witte & Dino Di Carlo
PNAS, March 2024
Abstract
The ability to selectively bind to antigenic peptides and secrete effector molecules can define rare and low-affinity populations of cells with therapeutic potential in emerging T cell receptor (TCR) immunotherapies. We leverage cavity-containing hydrogel microparticles, called nanovials, each coated with peptide-major histocompatibility complex (pMHC) monomers to isolate antigen-reactive T cells. T cells are captured and activated by pMHCs inducing the secretion of effector molecules including IFN-γ and granzyme B that are accumulated on nanovials, allowing sorting based on both binding and function. The TCRs of sorted cells on nanovials are sequenced, recovering paired αβ-chains using microfluidic emulsion-based single-cell sequencing. By labeling nanovials having different pMHCs with unique oligonucleotide-barcodes and secretions with oligo-barcoded detection antibodies, we could accurately link TCR sequences to specific targets and rank each TCR based on the corresponding cell’s secretion level. Using the technique, we identified an expanded repertoire of functional TCRs targeting viral antigens with high specificity and found rare TCRs with activity against cancer-specific splicing-enhanced epitopes.
Figure 1. Overview of high-throughput analysis and isolation of antigen-specific T cells followed by recovery of a single-cell TCR library.
Topics
TCR Discovery, Cytokine Profiling
Cell Types
Human T Cells, Human PBMCs
Instruments
SONY SH800S, 10x Genomics Chromium