title: Development of a luciferase immunoprecipitation system assay to detect IgG antibodies against human respiratory syncytial virus nucleoprotein.
abstract: The nucleoprotein of respiratory syncytial virus (RSV-N) is immunogenic and elicits an IgG response following infection. The RSV-N gene was cloned into a mammalian expression vector, pREN2, and the expressed luciferase-tagged protein (Ruc-N) detected anti-RSV-N-specific IgG antibodies using a high-throughput immunoprecipitation method (the luciferase immunoprecipitation system [LIPS]-N(RSV) assay). The specificity of the assay was evaluated using monoclonal antibodies (MAbs) and monospecific pre- and postimmunization rabbit antisera. Blood serum samples from chimpanzees and humans with proven/probable RSV infection were also tested. The pre- and postimmunization serum samples from rabbits given human metapneumovirus (HMPV) or measles virus were negative when tested by the LIPS-N(RSV) assay, while antisera obtained after immunization with either the RSV-A or RSV-B strain gave positive signals in a dose-dependent manner. RSV-N MAb 858-3 gave a positive signal in the LIPS-N(RSV) assay, while MAbs against other paramyxovirus nucleoproteins or RSV-F or RSV-G did not. Serum samples from chimpanzees simultaneously immunized with vaccinia-RSV-F and vaccinia-RSV-G recombinant viruses were negative in the LIPS-N(RSV) assay; however, anti-RSV-N IgG responses were detected following subsequent RSV challenge. Seven of the 12 infants who were seronegative at 9 months of age had detectable anti-RSV-N antibodies when they were retested at 15 to 18 months of age. The LIPS-N(RSV) assay detects specific anti-RSV-N IgG responses that may be used as a biomarker of RSV infection.
authors.affiliation: Laboratory of Pediatric and Respiratory Virus Diseases, Division of Viral Products, Office of Vaccines Research and Review, CBER, FDA, Bethesda, Maryland, USA.
authors.forename: Sangeeta
authors.initials: S
authors.lastname: Kumari
authors.forename: Roberta Lynne
authors.initials: RL
authors.lastname: Crim
authors.forename: Ashwin
authors.initials: A
authors.lastname: Kulkarni
authors.forename: Susette A
authors.initials: SA
authors.lastname: Audet
authors.forename: Thembi
authors.initials: T
authors.lastname: Mdluli
authors.forename: Haruhiko
authors.initials: H
authors.lastname: Murata
authors.forename: Judy A
authors.initials: JA
authors.lastname: Beeler
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood*; D047468:Immunoprecipitation / Q000379:methods*; D007223:Infant; D008156:Luciferases / Q000032:analysis*; D009698:Nucleoproteins / Q000276:immunology*; D002679:Pan troglodytes; D011817:Rabbits; D018357:Respiratory Syncytial Virus Infections / Q000175:diagnosis*; D018113:Respiratory Syncytial Virus, Human / Q000276:immunology*; D012680:Sensitivity and Specificity; D014764:Viral Proteins / Q000276:immunology
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G; D009698:Nucleoproteins; D014764:Viral Proteins; D008156:Luciferases