title: Flexible label-free quantitative assay for antibodies to influenza virus hemagglutinins.
abstract: During the initial pandemic influenza H1N1 virus outbreak, assays such as hemagglutination inhibition and microneutralization provided important information on the relative protection afforded by the population's cross-reactivity from prior infections and immunizations with seasonal vaccines. However, these assays continue to be limited in that they are difficult to automate for high throughput, such as in pandemic situations, as well as to standardize between labs. Thus, new technologies are being sought to improve standardization, reliability, and throughput by using chemically defined reagents rather than whole cells and virions. We now report the use of a cell-free and label-free flu antibody biosensor assay (f-AbBA) for influenza research and diagnostics that utilizes recombinant hemagglutinin (HA) in conjunction with label-free biolayer interferometry technology to measure biomolecular interactions between the HA and specific anti-HA antibodies or sialylated ligands. We evaluated f-AbBA to determine anti-HA antibody binding activity in serum or plasma to assess vaccine-induced humoral responses. This assay can reveal the impact of antigenic difference on antibody binding to HA and also measure binding to different subtypes of HA. We also show that the biosensor assay can measure the ability of HA to bind a model sialylated receptor-like ligand. f-AbBA could be used in global surveillance laboratories since preliminary tests on desiccated HA probes showed no loss of activity after >2 months in storage at room temperature, indicating that the same reagent lots could be used in different laboratories to minimize interlaboratory assay fluctuation. Future development of such reagents and similar technologies may offer a robust platform for future influenza surveillance activities.
authors.affiliation: Influenza Division, NCIRD, OID, Centers for Disease Control and Prevention, Atlanta, GA 30333, USA.
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Carney
authors.forename: Aleksandr S
authors.initials: AS
authors.lastname: Lipatov
authors.forename: Arnold S
authors.initials: AS
authors.lastname: Monto
authors.forename: Ruben O
authors.initials: RO
authors.lastname: Donis
authors.forename: James
authors.initials: J
authors.lastname: Stevens
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D005289:Ferrets; D006389:Hemagglutinins, Viral; D053118:Influenza A Virus, H1N1 Subtype / Q000276:immunology; D053122:Influenza A Virus, H3N2 Subtype / Q000276:immunology; D053124:Influenza A Virus, H5N1 Subtype / Q000276:immunology; D007252:Influenza Vaccines / Q000276:immunology*; D007368:Interferometry / Q000379:methods*; D008297:Male; D011994:Recombinant Proteins; D014773:Virology / Q000379:methods*
chemical_list: D000914:Antibodies, Viral; D006389:Hemagglutinins, Viral; D007252:Influenza Vaccines; D011994:Recombinant Proteins; C458881:hemagglutinin fusogenic peptide, influenza virus