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.
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.
['Pacini|Giulia|G|', 'Ieronymaki|Matthaia|M|', 'Nuti|Francesca|F|', 'Sabatino|Giuseppina|G|', 'Larregola|Maud|M|', 'Aharoni|Rina|R|', 'Papini|Anna Maria|AM|', 'Rovero|Paolo|P|http://orcid.org/0000-0001-9577-5228']
D000818:Animals D001323:Autoantibodies D001402:B-Lymphocytes D003001:Cloning, Molecular D004681:Encephalomyelitis, Autoimmune, Experimental D018604:Epitope Mapping D000939:Epitopes D004926:Escherichia coli D005260:Female D015870:Gene Expression D006801:Humans D007106:Immune Sera D051379:Mice D008810:Mice, Inbred C57BL D008872:Microwaves D008958:Models, Molecular D009103:Multiple Sclerosis D063308:Myelin-Oligodendrocyte Glycoprotein D010455:Peptides D011994:Recombinant Proteins D060327:Solid-Phase Synthesis Techniques
epitope mapping antibodies myelin oligodendrocyte glycoprotein experimental autoimmune encephalomyelitis enzyme-linked immunosorbent assay microwave-assisted solid-phase synthesis