title: Competition-based cellular peptide binding assays for 13 prevalent HLA class I alleles using fluorescein-labeled synthetic peptides.
abstract: We report the development, validation, and application of competition-based peptide binding assays for 13 prevalent human leukocyte antigen (HLA) class I alleles. The assays are based on peptide binding to HLA molecules on living cells carrying the particular allele. Competition for binding between the test peptide of interest and a fluorescein-labeled HLA class I binding peptide is used as read out. The use of cell membrane-bound HLA class I molecules circumvents the need for laborious biochemical purification of these molecules in soluble form. Previously, we have applied this principle for HLA-A2 and HLA-A3. We now describe the assays for HLA-A1, HLA-A11, HLA-A24, HLA-A68, HLA-B7, HLA-B8, HLA-B14, HLA-B35, HLA-B60, HLA-B61, and HLA-B62. Together with HLA-A2 and HLA-A3, these alleles cover more than 95% of the Caucasian population. Several allele-specific parameters were determined for each assay. Using these assays, we identified novel HLA class I high-affinity binding peptides from HIVpol, p53, PRAME, and minor histocompatibility antigen HA-1. Thus these convenient and accurate peptide-binding assays will be useful for the identification of putative cytotoxic T lymphocyte epitopes presented on a diverse array of HLA class I molecules.
authors.affiliation: Department of Immunohematology and Blood Transfusion, Leiden University Medical Center, The Netherlands.
authors.forename: Jan H
authors.initials: JH
authors.lastname: Kessler
authors.forename: Bregje
authors.initials: B
authors.lastname: Mommaas
authors.forename: Tuna
authors.initials: T
authors.lastname: Mutis
authors.forename: Ivo
authors.initials: I
authors.lastname: Huijbers
authors.forename: Debby
authors.initials: D
authors.lastname: Vissers
authors.forename: Willemien E
authors.initials: WE
authors.lastname: Benckhuijsen
authors.forename: Geziena M Th
authors.initials: GM
authors.lastname: Schreuder
authors.forename: Rienk
authors.initials: R
authors.lastname: Offringa
authors.forename: Els
authors.initials: E
authors.lastname: Goulmy
authors.forename: Cornelis J M
authors.initials: CJ
authors.lastname: Melief
authors.forename: Sjoerd H
authors.initials: SH
authors.lastname: van der Burg
authors.forename: Jan W
authors.initials: JW
authors.lastname: Drijfhout
mesh_terms: D000483:Alleles; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000951:Antigens, Neoplasm / Q000276:immunology / Q000378:metabolism; D001667:Binding, Competitive; D002461:Cell Line, Transformed; D018984:Epitopes, T-Lymphocyte / Q000276:immunology / Q000378:metabolism; D015684:Gene Products, pol / Q000276:immunology / Q000378:metabolism; D005805:Genes, MHC Class I; D015488:HIV Antigens / Q000276:immunology / Q000378:metabolism; D015234:HLA-A Antigens / Q000276:immunology / Q000378:metabolism; D015235:HLA-B Antigens / Q000276:immunology / Q000378:metabolism; D015395:Histocompatibility Antigens Class I / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D020128:Inhibitory Concentration 50; D015778:Minor Histocompatibility Antigens / Q000276:immunology / Q000378:metabolism; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000276:immunology / Q000378:metabolism; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000276:immunology* / Q000378:metabolism; D011485:Protein Binding; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D016159:Tumor Suppressor Protein p53 / Q000276:immunology / Q000378:metabolism
chemical_list: D000951:Antigens, Neoplasm; D018984:Epitopes, T-Lymphocyte; D015684:Gene Products, pol; C110615:HA-1 antigen; D015488:HIV Antigens; D015234:HLA-A Antigens; D015235:HLA-B Antigens; D015395:Histocompatibility Antigens Class I; D015778:Minor Histocompatibility Antigens; D009842:Oligopeptides; C404944:PRAME protein, human; D010446:Peptide Fragments; D016159:Tumor Suppressor Protein p53