title: Development of a simple, rapid, sensitive, high-throughput luciferase reporter based microneutralization test for measurement of virus neutralizing antibodies following Respiratory Syncytial Virus vaccination and infection.
abstract: We have established a new reporter gene-based RSV neutralization test using Renilla luciferase. The RSV-Luciferase Neutralization Test (RSV-Luc-NeuT) is a simple, rapid, high throughput, and less labor intensive functional serological assay than the traditional RSV-PRNT, capable of measuring a broad range of anti-RSV neutralizing antibodies targeting both RSV-F and RSV-G proteins. Specificity and sensitivity of the RSV-Luc-NeuT are comparable to the RSV-PRNT. Panels of pre-vaccination and post-vaccination animal sera, monoclonal antibodies and animal polyclonal anti-RSV sera confirmed assay specificity. A panel of 60 human sera demonstrated high assay sensitivity for measurement of RSV neutralizing antibodies that strongly correlated with the RSV-PRNT titers (R(2)=0.864). Neutralization in the presence of guinea pig complement (GPC) increased PRNT titers more than the RSV-Luc-NeuT neutralizing antibody titers for these human sera. This newly developed simple, high throughput, RSV-Luc-NeuT could be easily automated and applied in measurement of RSV neutralization titers in large vaccine trials.
authors.affiliation: Division of Viral Products, Center for Biologics Evaluation and Research (CBER), FDA, Bethesda, MD 20892, USA.
authors.forename: Sandra
authors.initials: S
authors.lastname: Fuentes
authors.forename: Roberta L
authors.initials: RL
authors.lastname: Crim
authors.forename: Judy
authors.initials: J
authors.lastname: Beeler
authors.forename: Michael N
authors.initials: MN
authors.lastname: Teng
authors.forename: Hana
authors.initials: H
authors.lastname: Golding
authors.forename: Surender
authors.initials: S
authors.lastname: Khurana
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000032:analysis; D002460:Cell Line / Q000821:virology; D003165:Complement System Proteins / Q000276:immunology; D000939:Epitopes / Q000276:immunology; D017930:Genes, Reporter; D057166:High-Throughput Screening Assays; D006801:Humans; D007106:Immune Sera; D008156:Luciferases / Q000235:genetics*; D051379:Mice; D009500:Neutralization Tests / Q000379:methods*; D011817:Rabbits; D012136:Respiratory Syncytial Viruses / Q000276:immunology*; D012680:Sensitivity and Specificity; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology; D014760:Viral Fusion Proteins / Q000276:immunology
chemical_list: D057134:Antibodies, Neutralizing; D000939:Epitopes; C526850:F protein, human respiratory syncytial virus; D007106:Immune Sera; D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins; C116204:attachment protein G; D003165:Complement System Proteins; D008156:Luciferases
keywords: Antibody; Assay; F protein; G protein; Infection; Luciferase; Neutralization; PRNT; RSV; RSV-Luc-NeuT; Rapid; Respiratory Syncytial Virus; Vaccine