title: Disulfide-modified antigen for detection of celiac disease-associated anti-tissue transglutaminase autoantibodies.
abstract: A simple and rapid immunosensor for the determination of the celiac disease-related antibody, anti-tissue transglutaminase, was investigated. The antigenic protein tissue transglutaminase was chemically modified, introducing disulfide groups through different moieties of the molecule (amine, carboxylic, and hydroxyl groups), self-assembled on gold surfaces, and used for the detection of IgA and IgG autoantibodies. The modified proteins were evaluated using enzyme-linked immunosorbent assay and surface plasmon resonance, which showed that only introduction of the disulfide groups through amine moieties in the tissue transglutaminase preserved its antigenic properties. The disulfide-modified antigen was co-immobilized via chemisorption with a poly(ethylene glycol) alkanethiol on gold electrodes. The modified electrodes were then exposed to IgA anti-tissue transglutaminase antibodies and subsequently to horseradish peroxidase-labeled anti-idiotypic antibodies, achieving a detection limit of 260 ng ml-1. Immunosensor performance in the presence of complex matrixes, including clinically relevant serum reference solutions and real patient samples, was evaluated. The introduction of disulfides in the antigenic protein enabled a simple and convenient one-step surface immobilization procedure involving only spontaneous gold-thiol covalent binding. Complete amperometric assay time was 30 min.
authors.affiliation: Universitat Rovira i Virgili, Avinguda Països Catalans 26, 43007, Tarragona, Spain.|Departamento de Ingeniería Química, Universidad de Guadalajara, Guadalajara, México.
authors.forename: Luis Carlos
authors.initials: LC
authors.lastname: Rosales-Rivera
authors.affiliation: Universitat Rovira i Virgili, Avinguda Països Catalans 26, 43007, Tarragona, Spain.
authors.forename: Samuel
authors.initials: S
authors.lastname: Dulay
authors.affiliation: Universitat Rovira i Virgili, Avinguda Països Catalans 26, 43007, Tarragona, Spain.|Integrated Microsystems for Quality of Life, s.l., 43006 Pol. Ind. Riu Clar, Tarragona, Spain.
authors.forename: Pablo
authors.initials: P
authors.lastname: Lozano-Sánchez
authors.affiliation: Universitat Rovira i Virgili, Avinguda Països Catalans 26, 43007, Tarragona, Spain.
authors.forename: Ioanis
authors.initials: I
authors.lastname: Katakis
authors.affiliation: Universitat Rovira i Virgili, Avinguda Països Catalans 26, 43007, Tarragona, Spain. joseplluis.acero@urv.cat.
authors.forename: Josep Lluís
authors.initials: JL
authors.lastname: Acero-Sánchez
authors.affiliation: Universitat Rovira i Virgili, Avinguda Països Catalans 26, 43007, Tarragona, Spain. ciara.osullivan@urv.cat.|Institució Catalana de Recerca i Estudis Avançats, Passeig Lluís Companys, 23, 08010, Barcelona, Spain. ciara.osullivan@urv.cat.
authors.forename: Ciara K
authors.initials: CK
authors.lastname: O'Sullivan
mesh_terms: D001323:Autoantibodies / Q000032:analysis* / Q000097:blood / Q000276:immunology; D015374:Biosensing Techniques / Q000379:methods*; D002446:Celiac Disease / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D004220:Disulfides / Q000737:chemistry* / Q000276:immunology; D055664:Electrochemical Techniques / Q000379:methods; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000276:immunology; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000276:immunology; D006801:Humans; D007118:Immunoassay / Q000379:methods; D007070:Immunoglobulin A / Q000032:analysis / Q000097:blood / Q000276:immunology; D007074:Immunoglobulin G / Q000032:analysis / Q000097:blood / Q000276:immunology; D057230:Limit of Detection; D008958:Models, Molecular; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases / Q000737:chemistry* / Q000276:immunology
chemical_list: D001323:Autoantibodies; D004220:Disulfides; D004800:Enzymes, Immobilized; D007070:Immunoglobulin A; D007074:Immunoglobulin G; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D019204:GTP-Binding Proteins
keywords: Anti-tissue transglutaminase antibodies; Celiac disease; Electrochemical immunosensor; Modified proteins