High-throughput multi-dimensional single antigen-specific T cell profiling

Abstract

Abstract Recent advances in single cell analysis has made it possible to interrogate the transcriptome of single cells in high-throughput manner. However, multi-dimensional profiling of antigen-specific T cells at single cell level for hundreds of antigens has not been realized, which becomes a bottleneck to the understanding of these important cells in immunological diseases. Here we describe a new technology, TetTCR-SeqHD, that extends our previously developed technology, TetTCR-Seq for Tetramer associated TCR sequencing, to additional two dimensions, single cell gene expression and phenotype. In addition, we expand the throughput to tens of thousands of single T cells. We achieved these by using DNA-barcoded pMHC tetramers and antibodies in combination with single cell gene expression analysis, thus, paralleling all read-outs using next-gen sequencing. To validate the technology, we have used previously generated polyclonal T cells to 6 different antigens with known TCR sequences. This analysis showed that we could achieve less than 5% on false discovery rate. We then used TetTCR-SeqHD on a panel of type 1 diabetes related peptides and showed unexpected features of antigen-specific T cells that separate healthy individuals from type 1 diabetes patients. In addition, TCR transgenics T cells confirmed their antigen specificity. In summary, TetTCR-SeqHD enables us to analyze antigen-specific T cells with an extraordinary depth and breadth, which enables discoveries directly in humans. TetTCR-SeqHD would be a useful technology for studying many different diseases where screening for T cells antigens and deep profiling of antigen-specific T cells are highly sought after.