title: Enhanced Detection of Antigen-Specific CD4+ T Cells Using Altered Peptide Flanking Residue Peptide-MHC Class II Multimers.
abstract: Fluorochrome-conjugated peptide-MHC (pMHC) class I multimers are staple components of the immunologist's toolbox, enabling reliable quantification and analysis of Ag-specific CD8(+) T cells irrespective of functional outputs. In contrast, widespread use of the equivalent pMHC class II (pMHC-II) reagents has been hindered by intrinsically weaker TCR affinities for pMHC-II, a lack of cooperative binding between the TCR and CD4 coreceptor, and a low frequency of Ag-specific CD4(+) T cell populations in the peripheral blood. In this study, we show that peptide flanking regions, extending beyond the central nonamer core of MHC-II-bound peptides, can enhance TCR-pMHC-II binding and T cell activation without loss of specificity. Consistent with these findings, pMHC-II multimers incorporating peptide flanking residue modifications proved superior for the ex vivo detection, characterization, and manipulation of Ag-specific CD4(+) T cells, highlighting an unappreciated feature of TCR-pMHC-II interactions.
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Holland
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and.
authors.forename: Garry
authors.identifier: http://orcid.org/0000-0002-6332-9108
authors.initials: G
authors.lastname: Dolton
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and.
authors.forename: Martin
authors.initials: M
authors.lastname: Scurr
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and.
authors.forename: Kristin
authors.identifier: http://orcid.org/0000-0002-9856-2938
authors.initials: K
authors.lastname: Ladell
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and.
authors.forename: Andrea J
authors.initials: AJ
authors.lastname: Schauenburg
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and.
authors.forename: Kelly
authors.initials: K
authors.lastname: Miners
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and.
authors.forename: Florian
authors.initials: F
authors.lastname: Madura
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and.
authors.forename: Andrew K
authors.initials: AK
authors.lastname: Sewell
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and.
authors.forename: David A
authors.initials: DA
authors.lastname: Price
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and godkinaj@cardiff.ac.uk coledk@cardiff.ac.uk.
authors.forename: David K
authors.initials: DK
authors.lastname: Cole
authors.affiliation: Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, Wales, United Kingdom; and Department of Integrated Medicine, University Hospital of Wales, Cardiff CF14 4XW, Wales, United Kingdom godkinaj@cardiff.ac.uk coledk@cardiff.ac.uk.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Godkin
mesh_terms: D000941:Antigens / Q000378:metabolism*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D002460:Cell Line; D002469:Cell Separation / Q000379:methods*; D006680:HLA Antigens / Q000276:immunology*; D000949:Histocompatibility Antigens Class II / Q000378:metabolism; D006801:Humans; D008213:Lymphocyte Activation; D010446:Peptide Fragments / Q000378:metabolism*; D011485:Protein Binding; D011948:Receptors, Antigen, T-Cell / Q000276:immunology; D037182:T-Cell Antigen Receptor Specificity
chemical_list: D000941:Antigens; D006680:HLA Antigens; D000949:Histocompatibility Antigens Class II; D010446:Peptide Fragments; D011948:Receptors, Antigen, T-Cell