title: A novel assay for detecting virus-specific antibodies triggering activation of Fcγ receptors.
abstract: IgG responses are crucial in antiviral defence and instrumental for the serodiagnosis of infections. Fcγ receptors (FcγRs), which recognize the Fc-part of IgG, differ regarding their IgG binding affinity, IgG subclass preference, cellular expression profile and pathogen elimination mechanisms elicited upon activation. Assessing their activation in vitro is of fundamental importance, but technically difficult. Therefore, a novel assay for measuring antiviral IgG antibodies triggering activation of individual host Fcγ receptors was established. The assay comprises the co-cultivation of virus-infected target cells with immune IgG antibodies and mouse BW5147 hybridoma cells stably expressing chimeric FcγR-CD3ζ chain molecules consisting of the extracellular domain of human FcγRIIIA, FcγRIIA or FcγRI, respectively, fused to the transmembrane and intracellular domains of the mouse CD3ζ chain. Triggering of the chimeric FcγR receptors by immune complexes formed on the surface of IgG-opsonized virus-infected target cells resulted in FcγR activation leading to IL-2 secretion by BW5147 cells, which was quantified as a surrogate marker in an ELISA. Target cells infected with various human pathogenic viruses including herpes simplex virus type 1 (HSV-1), Epstein-Barr virus (EBV), human cytomegalovirus (HCMV), measles virus (MV), and respiratory syncytial virus (RSV) or displaying human immunodeficiency virus-1 (HIV-1) gp120 evoke dose-dependent IgG responses demonstrating the universal applicability of the assay. Taken together, a new reliable and simple tool for measuring antibodies triggering activation of Fcγ receptors was established. This assay will be instrumental for defining novel correlates of IgG immunity and the design of new therapeutic IgGs.
authors.affiliation: Institute for Virology, Heinrich-Heine-University Düsseldorf, Universitätsstrasse 1, 40225 Düsseldorf, Germany.
authors.forename: Eugenia
authors.initials: E
authors.lastname: Corrales-Aguilar
authors.forename: Mirko
authors.initials: M
authors.lastname: Trilling
authors.forename: Henrike
authors.initials: H
authors.lastname: Reinhard
authors.forename: Eva
authors.initials: E
authors.lastname: Mercé-Maldonado
authors.forename: Marek
authors.initials: M
authors.lastname: Widera
authors.forename: Heiner
authors.initials: H
authors.lastname: Schaal
authors.forename: Albert
authors.initials: A
authors.lastname: Zimmermann
authors.forename: Ofer
authors.initials: O
authors.lastname: Mandelboim
authors.forename: Hartmut
authors.initials: H
authors.lastname: Hengel
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology* / Q000378:metabolism; D017252:CD3 Complex / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D002460:Cell Line; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D018920:Coculture Techniques; D003587:Cytomegalovirus / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D057809:HEK293 Cells; D018259:Herpesvirus 1, Human / Q000276:immunology; D004854:Herpesvirus 4, Human / Q000276:immunology; D006801:Humans; D006825:Hybridomas; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism; D007376:Interleukin-2 / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology; D007694:Killer Cells, Natural / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D051379:Mice; D011485:Protein Binding / Q000276:immunology; D017452:Receptors, IgG / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D020298:U937 Cells; D014709:Vero Cells; D014780:Viruses / Q000276:immunology*
chemical_list: D000914:Antibodies, Viral; D017252:CD3 Complex; C121704:CD3 antigen, zeta chain; D007074:Immunoglobulin G; D007376:Interleukin-2; D017452:Receptors, IgG; D011993:Recombinant Fusion Proteins