title: T cells recognizing a peptide contaminant undetectable by mass spectrometry.
abstract: Synthetic peptides are widely used in immunological research as epitopes to stimulate their cognate T cells. These preparations are never completely pure, but trace contaminants are commonly revealed by mass spectrometry quality controls. In an effort to characterize novel major histocompatibility complex (MHC) Class I-restricted β-cell epitopes in non-obese diabetic (NOD) mice, we identified islet-infiltrating CD8+ T cells recognizing a contaminating peptide. The amount of this contaminant was so small to be undetectable by direct mass spectrometry. Only after concentration by liquid chromatography, we observed a mass peak corresponding to an immunodominant islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)(206-214) epitope described in the literature. Generation of CD8+ T-cell clones recognizing IGRP(206-214) using a novel method confirmed the identity of the contaminant, further underlining the immunodominance of IGRP(206-214). If left undetected, minute impurities in synthetic peptide preparations may thus give spurious results.
authors.affiliation: INSERM, U986, DeAR Lab Avenir, Saint Vincent de Paul Hospital, Paris, France.
authors.forename: Vedran
authors.initials: V
authors.lastname: Brezar
authors.forename: Slobodan
authors.initials: S
authors.lastname: Culina
authors.forename: Thomas
authors.initials: T
authors.lastname: Østerbye
authors.forename: François
authors.initials: F
authors.lastname: Guillonneau
authors.forename: Giovanni
authors.initials: G
authors.lastname: Chiappetta
authors.forename: Yann
authors.initials: Y
authors.lastname: Verdier
authors.forename: Joelle
authors.initials: J
authors.lastname: Vinh
authors.forename: F Susan
authors.initials: FS
authors.lastname: Wong
authors.forename: Søren
authors.initials: S
authors.lastname: Buus
authors.forename: Roberto
authors.initials: R
authors.lastname: Mallone
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D002999:Clone Cells; D018984:Epitopes, T-Lymphocyte / Q000276:immunology; D005952:Glucose-6-Phosphatase / Q000276:immunology; D015395:Histocompatibility Antigens Class I / Q000276:immunology; D006801:Humans; D007371:Interferon-gamma / Q000276:immunology; D007515:Islets of Langerhans / Q000276:immunology; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D016688:Mice, Inbred NOD; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000276:immunology*; D011506:Proteins / Q000276:immunology; D015203:Reproducibility of Results; D020539:Sequence Analysis, Protein
chemical_list: D018984:Epitopes, T-Lymphocyte; D015395:Histocompatibility Antigens Class I; D010455:Peptides; D011506:Proteins; D007371:Interferon-gamma; D005952:Glucose-6-Phosphatase; C117983:G6pc2 protein, mouse