title: Profiling Humoral Immune Responses to Clostridium difficile-Specific Antigens by Protein Microarray Analysis.
abstract: Clostridium difficile is an anaerobic, Gram-positive, and spore-forming bacterium that is the leading worldwide infective cause of hospital-acquired and antibiotic-associated diarrhea. Several studies have reported associations between humoral immunity and the clinical course of C. difficile infection (CDI). Host humoral immune responses are determined using conventional enzyme-linked immunosorbent assay (ELISA) techniques. Herein, we report the first use of a novel protein microarray assay to determine systemic IgG antibody responses against a panel of highly purified C. difficile-specific antigens, including native toxins A and B (TcdA and TcdB, respectively), recombinant fragments of toxins A and B (TxA4 and TxB4, respectively), ribotype-specific surface layer proteins (SLPs; 001, 002, 027), and control proteins (tetanus toxoid and Candida albicans). Microarrays were probed with sera from a total of 327 individuals with CDI, cystic fibrosis without diarrhea, and healthy controls. For all antigens, precision profiles demonstrated <10% coefficient of variation (CV). Significant correlation was observed between microarray and ELISA in the quantification of antitoxin A and antitoxin B IgG. These results indicate that microarray is a suitable assay for defining humoral immune responses to C. difficile protein antigens and may have potential advantages in throughput, convenience, and cost.
authors.affiliation: Immunology, School of Life Sciences, University of Nottingham, Nottingham, United Kingdom Medical Microbiology and Immunology, Faculty of Medicine, Mansoura University, Mansoura, Egypt.
authors.forename: Ola H
authors.initials: OH
authors.lastname: Negm
authors.affiliation: Immunology, School of Life Sciences, University of Nottingham, Nottingham, United Kingdom Medical Microbiology and Immunology, Faculty of Medicine, Mansoura University, Mansoura, Egypt.
authors.forename: Mohamed R
authors.initials: MR
authors.lastname: Hamed
authors.affiliation: Immunology, School of Life Sciences, University of Nottingham, Nottingham, United Kingdom.
authors.forename: Elizabeth M
authors.initials: EM
authors.lastname: Dilnot
authors.affiliation: Public Health England, Salisbury, United Kingdom.
authors.forename: Clifford C
authors.initials: CC
authors.lastname: Shone
authors.affiliation: Immunomodulation Research Group, Dublin City University, Dublin, Ireland.
authors.forename: Izabela
authors.initials: I
authors.lastname: Marszalowska
authors.affiliation: Immunomodulation Research Group, Dublin City University, Dublin, Ireland.
authors.forename: Mark
authors.initials: M
authors.lastname: Lynch
authors.affiliation: Immunomodulation Research Group, Dublin City University, Dublin, Ireland.
authors.forename: Christine E
authors.initials: CE
authors.lastname: Loscher
authors.affiliation: Faculty of Medicine and Health Sciences, University of Nottingham, Nottingham, United Kingdom.
authors.forename: Laura J
authors.initials: LJ
authors.lastname: Edwards
authors.affiliation: Immunology, School of Life Sciences, University of Nottingham, Nottingham, United Kingdom.
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Tighe
authors.affiliation: Leeds Institute for Molecular Medicine, University of Leeds, Leeds, United Kingdom.
authors.forename: Mark H
authors.initials: MH
authors.lastname: Wilcox
authors.affiliation: NIHR Nottingham Digestive Diseases Biomedical Research Unit, Nottingham, United Kingdom tanya.monaghan@nottingham.ac.uk.
authors.forename: Tanya M
authors.initials: TM
authors.lastname: Monaghan
mesh_terms: D000328:Adult; D000368:Aged; D000942:Antigens, Bacterial / Q000276:immunology*; D001426:Bacterial Proteins / Q000276:immunology; D001427:Bacterial Toxins / Q000276:immunology; D016360:Clostridioides difficile / Q000276:immunology*; D004768:Enterotoxins / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D006801:Humans; D056724:Immunity, Humoral / Q000276:immunology*; D007074:Immunoglobulin G / Q000097:blood; D008297:Male; D008875:Middle Aged; D040081:Protein Array Analysis / Q000379:methods*; D037521:Virulence Factors / Q000276:immunology; D055815:Young Adult
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004768:Enterotoxins; D007074:Immunoglobulin G; D037521:Virulence Factors; C041348:tcdA protein, Clostridium difficile; C031815:toxB protein, Clostridium difficile