title: Epitope mapping of anti-myelin oligodendrocyte glycoprotein (MOG) antibodies in a mouse model of multiple sclerosis: microwave-assisted synthesis of the peptide antigens and ELISA screening.
abstract: The role of pathologic auto-antibodies against myelin oligodendrocyte glycoprotein (MOG) in multiple sclerosis is a highly controversial matter. As the use of animal models may enable to unravel the molecular mechanisms of the human disorder, numerous studies on multiple sclerosis are carried out using experimental autoimmune encephalomyelitis (EAE). In particular, the most extensively used EAE model is obtained by immunizing C57BL/6 mice with the immunodominant peptide MOG(35-55). In this scenario, we analyzed the anti-MOG antibody response in this model using the recombinant refolded extracellular domain of the protein, MOG(1-117). To assess the presence of a B-cell intramolecular epitope spreading mechanism, we tested also five synthetic peptides mapping the 1-117 sequence of MOG, including MOG(35-55). For this purpose, we cloned, expressed in Escherichia coli and on-column refolded MOG(1-117), and we applied an optimized microwave-assisted solid-phase synthetic strategy to obtain the designed peptide sequences. Subsequently, we set up a solid-phase immunoenzymatic assay testing both naïve and EAE mice sera and using MOG protein and peptides as antigenic probes. The results obtained disclose an intense IgG antibody response against both the recombinant protein and the immunizing peptide, while no response was observed against the other synthetic fragments, thus excluding the presence of an intramolecular epitope spreading mechanism. Furthermore, as the properly refolded recombinant probe is able to bind antibodies with greater efficiency compared with MOG(35-55), we hypothesize the presence of both linear and conformational epitopes on MOG(35-55) sequence.
authors.affiliation: French-Italian Interdepartmental Laboratory of Peptide and Protein Chemistry and Biology - PeptLab, Florence, Italy and Cergy-Pontoise, France.|Department NeuroFarBa, Section of Pharmaceutical and Nutraceutical Sciences, University of Florence, Via Ugo Schiff 6, Sesto Fiorentino, Florence, I-50019, Italy.
authors.forename: Giulia
authors.initials: G
authors.lastname: Pacini
authors.affiliation: French-Italian Interdepartmental Laboratory of Peptide and Protein Chemistry and Biology - PeptLab, Florence, Italy and Cergy-Pontoise, France.|Department NeuroFarBa, Section of Pharmaceutical and Nutraceutical Sciences, University of Florence, Via Ugo Schiff 6, Sesto Fiorentino, Florence, I-50019, Italy.|Laboratoire de Chimie Biologique EA4505, University of Cergy-Pontoise, 5 mail Gay-Lussac Neuville-sur-Oise, Cergy-Pontoise, 95000, France.
authors.forename: Matthaia
authors.initials: M
authors.lastname: Ieronymaki
authors.affiliation: French-Italian Interdepartmental Laboratory of Peptide and Protein Chemistry and Biology - PeptLab, Florence, Italy and Cergy-Pontoise, France.|Department of Chemistry "Ugo Schiff", University of Florence, Via della Lastruccia 3/13, Sesto Fiorentino, Florence, I-50019, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Nuti
authors.affiliation: French-Italian Interdepartmental Laboratory of Peptide and Protein Chemistry and Biology - PeptLab, Florence, Italy and Cergy-Pontoise, France.|Department of Chemistry "Ugo Schiff", University of Florence, Via della Lastruccia 3/13, Sesto Fiorentino, Florence, I-50019, Italy.
authors.forename: Giuseppina
authors.initials: G
authors.lastname: Sabatino
authors.affiliation: French-Italian Interdepartmental Laboratory of Peptide and Protein Chemistry and Biology - PeptLab, Florence, Italy and Cergy-Pontoise, France.|Laboratoire de Chimie Biologique EA4505, University of Cergy-Pontoise, 5 mail Gay-Lussac Neuville-sur-Oise, Cergy-Pontoise, 95000, France.
authors.forename: Maud
authors.initials: M
authors.lastname: Larregola
authors.affiliation: Department of Immunology, The Weizmann Institute of Science, Rehovot, 76100, Israel.
authors.forename: Rina
authors.initials: R
authors.lastname: Aharoni
authors.affiliation: French-Italian Interdepartmental Laboratory of Peptide and Protein Chemistry and Biology - PeptLab, Florence, Italy and Cergy-Pontoise, France.|Department of Chemistry "Ugo Schiff", University of Florence, Via della Lastruccia 3/13, Sesto Fiorentino, Florence, I-50019, Italy.|Laboratoire de Chimie Biologique EA4505, University of Cergy-Pontoise, 5 mail Gay-Lussac Neuville-sur-Oise, Cergy-Pontoise, 95000, France.
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Papini
authors.affiliation: French-Italian Interdepartmental Laboratory of Peptide and Protein Chemistry and Biology - PeptLab, Florence, Italy and Cergy-Pontoise, France.|Department NeuroFarBa, Section of Pharmaceutical and Nutraceutical Sciences, University of Florence, Via Ugo Schiff 6, Sesto Fiorentino, Florence, I-50019, Italy.
authors.forename: Paolo
authors.identifier: http://orcid.org/0000-0001-9577-5228
authors.initials: P
authors.lastname: Rovero
mesh_terms: D000818:Animals; D001323:Autoantibodies / Q000096:biosynthesis / Q000737:chemistry*; D001402:B-Lymphocytes / Q000276:immunology; D003001:Cloning, Molecular; D004681:Encephalomyelitis, Autoimmune, Experimental / Q000235:genetics / Q000276:immunology* / Q000473:pathology; D018604:Epitope Mapping; D000939:Epitopes / Q000737:chemistry* / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D006801:Humans; D007106:Immune Sera / Q000737:chemistry; D051379:Mice; D008810:Mice, Inbred C57BL; D008872:Microwaves; D008958:Models, Molecular; D009103:Multiple Sclerosis / Q000235:genetics / Q000276:immunology / Q000473:pathology; D063308:Myelin-Oligodendrocyte Glycoprotein / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D010455:Peptides / Q000138:chemical synthesis* / Q000276:immunology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D060327:Solid-Phase Synthesis Techniques / Q000379:methods
chemical_list: D001323:Autoantibodies; D000939:Epitopes; D007106:Immune Sera; D063308:Myelin-Oligodendrocyte Glycoprotein; D010455:Peptides; D011994:Recombinant Proteins
keywords: antibodies; enzyme-linked immunosorbent assay; epitope mapping; experimental autoimmune encephalomyelitis; microwave-assisted solid-phase synthesis; myelin oligodendrocyte glycoprotein