biostudies-literatureFahad ASUniversity of Kansas Cancer CenterUniversity of Minnesota FoundationNIAID NIH HHSRandy Shaver Cancer Research and Community FundNCI NIH HHSNational Institutes of HealthUS Department of DefenseWellcome TrustNIH HHSNIGMS NIH HHSRichard M. Schulze Family Foundationgzab034https://www.ebi.ac.uk/biostudies/studies/S-EPMC9005053biostudies-literatureUnknown35Functional analyses of the T cell receptor (TCR) landscape can reveal critical information about protection from disease and molecular responses to vaccines. However, it has proven difficult to combine advanced next-generation sequencing technologies with methods to decode the peptide-major histocompatibility complex (pMHC) specificity of individual TCRs. We developed a new high-throughput approach to enable repertoire-scale functional evaluations of natively paired TCRs. In particular, we leveraged the immortalized nature of physically linked TCRα:β amplicon libraries to analyze binding against multiple recombinant pMHCs on a repertoire scale, and to exemplify the utility of this approach, we also performed affinity-based functional mapping in conjunction with quantitative next-generationProtein engineering, design & selection : PEDSImmortalization and functional screening of natively paired human T cell receptor repertoires.PMC9005053P20 GM103418P20 GM103638100326/Z/12/ZDP5OD023118R21 AI166396W81XWH1810296R21 CA230487P20GM103638P20GM103418DP5 OD023118R21CA230487Douek DCLlewellyn-Lacey SPrice DALaflin ADFahad ASDeKosky BJMadan BBalfour HHMatus-Nicodemos RKoup RABoyle NWolfe JChung CYLopez Acevedo SNLadi RRGutierrez-Gonzalez MFZhou JfalseImmortalization and functional screening of natively paired human T cell receptor repertoires.Functional analyses of the T cell receptor (TCR) landscape can reveal critical information about protection from disease and molecular responses to vaccines. However, it has proven difficult to combine advanced next-generation sequencing technologies with methods to decode the peptide-major histocompatibility complex (pMHC) specificity of individual TCRs. We developed a new high-throughput approach to enable repertoire-scale functional evaluations of natively paired TCRs. In particular, we leveraged the immortalized nature of physically linked TCRα:β amplicon libraries to analyze binding against multiple recombinant pMHCs on a repertoire scale, and to exemplify the utility of this approach, we also performed affinity-based functional mapping in conjunction with quantitative next-generation2022-01-01T00:00:00Z2022 Feb2026-06-03T17:08:08.021Z2025-04-21T15:56:58.839ZS-EPMC90050533517485910.1093/protein/gzab034