title: Multiplexed infectious protein microarray immunoassay suitable for the study of the specificity of monoclonal immunoglobulins.
abstract: Enzyme-linked immunosorbent assays (ELISAs) used to detect antibodies specific for common infectious agents such as Epstein-Barr virus (EBV), cytomegalovirus (CMV), Toxoplasma gondii (T. gondii), and hepatitis C virus (HCV) are time-consuming and require large volumes of samples, which restrict their use. We propose a new assay based on a multiplexed infectious protein (MIP) microarray combining different epitopes representative of the four germs. Antigens and lysates were printed on nitrocellulose slides to constitute the microarray. First, the microarray was incubated with human serum samples. Then, the suitability of the microarray for analysis of the specificity of purified monoclonal immunoglobulin (mc Ig) was assessed using serum and mc Ig of HCV-positive patients. Bound human immunoglobulin G (IgG) was detected using fluorescently labeled secondary antibodies, and the signals were quantified. Results obtained in serum samples with the new MIP microarray immunoassay were compared with ELISAs; we observed concordances of 95% for EBV, 93% for CMV, 91% for T. gondii, and 100% for HCV. Regarding purified mc Ig of HCV-positive patients, 3 of 3 recognized antigens printed on the microarray. Hence, the novel EBV/CMV/T. gondii/HCV MIP microarray allows simultaneous diagnosis of polyclonal and monoclonal immune response to infectious diseases using very small volume samples.
authors.affiliation: INSERM UMR892/CNRS UMR6299, Institut de Recherche Thérapeutique, Université de Nantes, 44007 Nantes Cedex 1, France.
authors.forename: Delphine
authors.initials: D
authors.lastname: Feron
authors.forename: Cathy
authors.initials: C
authors.lastname: Charlier
authors.forename: Victor
authors.initials: V
authors.lastname: Gourain
authors.forename: Laurent
authors.initials: L
authors.lastname: Garderet
authors.forename: Marianne
authors.initials: M
authors.lastname: Coste-Burel
authors.forename: Patrice
authors.initials: P
authors.lastname: Le Pape
authors.forename: Pierre
authors.initials: P
authors.lastname: Weigel
authors.forename: Yannick
authors.initials: Y
authors.lastname: Jacques
authors.forename: Sylvie
authors.initials: S
authors.lastname: Hermouet
authors.forename: Edith
authors.initials: E
authors.lastname: Bigot-Corbel
mesh_terms: D000328:Adult; D000913:Antibodies, Protozoan / Q000097:blood* / Q000276:immunology; D000914:Antibodies, Viral / Q000097:blood* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect / Q000379:methods; D006801:Humans; D008297:Male; D008875:Middle Aged; D040081:Protein Array Analysis / Q000379:methods*; D012680:Sensitivity and Specificity; D014122:Toxoplasma; D014123:Toxoplasmosis / Q000097:blood* / Q000175:diagnosis / Q000276:immunology; D014777:Virus Diseases / Q000097:blood* / Q000175:diagnosis / Q000276:immunology; D014780:Viruses
chemical_list: D000913:Antibodies, Protozoan; D000914:Antibodies, Viral