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Identification of antigen-specific functional CD8+ T cells using an optofluidic system independent of epitope
Wei Zhang1, Yu Liu1, Hongxiang Zeng1
1The Second Affiliated Hospital, School of Medicine, Southern University of Science and Technology, Institute for Hepatology, National Clinical Research Center for Infectious Disease, Shenzhen Third People's Hospital, Shenzhen, Guangdong Province 518112, China.
Most studies detect and isolate antigen-specific CD8+ T cells solely by measuring T cell receptor (TCR) binding affinity to peptide-major histocompatibility complexes (p-MHCs) without considering T cell function. Here, we present a function-based strategy to screen antigen-specific CD8+ T cells using the Lightning optofluidic platform, named MicroFAST (microfluidic function-based screening of antigen-specific single T lymphocyte) fitting for limited sample volume. We validated this method by using it to isolate human cytomegalovirus (HCMV)-specific CD8+ T cells targeting a known HLA-A∗02:01-restricted epitope. We finally applied this method to identify several SARS-CoV-2-specific TCRs, which were characterized by recognizing HLA-B∗35:01-restricted TPS epitope (TPSGTWLTY), demonstrating the utility of our system. TCR binding affinity did not associate with its functionality. Collectively, we developed a workflow of isolating antigen-specific functional CD8+ T cells with or without a prior knowledge of peptide-MHC specificities, which will facilitate the development of engineered TCR-T therapies and T cell-based vaccines.
Most studies detect and isolate antigen-specific CD8+ T cells solely by measuring T cell receptor (TCR) binding affinity to peptide-major histocompatibility complexes (p-MHCs) without considering T cell function. Here, we present a function-based strategy to screen antigen-specific CD8+ T cells using the Lightning optofluidic platform, named MicroFAST (microfluidic function-based screening of antigen-specific single T lymphocyte) fitting for limited sample volume. We validated this method by using it to isolate human cytomegalovirus (HCMV)-specific CD8+ T cells targeting a known HLA-A∗02:01-restricted epitope. We finally applied this method to identify several SARS-CoV-2-specific TCRs, which were characterized by recognizing HLA-B∗35:01-restricted TPS epitope (TPSGTWLTY), demonstrating the utility of our system. TCR binding affinity did not associate with its functionality. Collectively, we developed a workflow of isolating antigen-specific functional CD8+ T cells with or without a prior knowledge of peptide-MHC specificities, which will facilitate the development of engineered TCR-T therapies and T cell-based vaccines.
