title: Higher Throughput Quantification of Neutralizing Antibody to Herpes Simplex Viruses.
abstract: We report a rapid, higher throughput method for measuring neutralizing antibody to herpes simplex virus (HSV) in human sera. Clinical isolates and sera from the Herpevac Trial for Women were used in a colorimetric assay in which infection of tissue culture (lack of neutralization) was indicated by substrate metabolism by beta-galactosidase induced in the ELVIS cell line. The neutralization assay was optimized by addition of guinea pig complement, which particularly enhanced neutralizing antibody titers to HSV-2. Higher neutralizing antibody titers were also achieved using virus particles isolated from the supernatant of infected cells rather than lysate of infected cells as the source of virus. The effect of assay incubation time and incubation time with substrate were also optimized. We found that incubating with substrate until a standard optical density of 1.0 was reached permitted a better comparison among virus isolates, and achieved reliable measurement of neutralizing antibody activity. Interestingly, in contrast to results in the absence of complement, addition of complement allowed sera from HSV-2 gD-vaccinated subjects to neutralize HSV-1 and HSV-2 clinical and laboratory isolates with equal potency.
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, Saint Louis University School of Medicine, St. Louis, Missouri, United States of America.
authors.forename: Tamara P
authors.initials: TP
authors.lastname: Blevins
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, Saint Louis University School of Medicine, St. Louis, Missouri, United States of America.
authors.forename: Michelle C
authors.initials: MC
authors.lastname: Mitchell
authors.affiliation: Department of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, St. Louis, Missouri, United States of America.
authors.forename: Maria
authors.initials: M
authors.lastname: Korom
authors.affiliation: Department of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, St. Louis, Missouri, United States of America.
authors.forename: Hong
authors.initials: H
authors.lastname: Wang
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, Saint Louis University School of Medicine, St. Louis, Missouri, United States of America.
authors.forename: Yinyi
authors.initials: Y
authors.lastname: Yu
authors.affiliation: Department of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, St. Louis, Missouri, United States of America.
authors.forename: Lynda A
authors.initials: LA
authors.lastname: Morrison
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, Saint Louis University School of Medicine, St. Louis, Missouri, United States of America.
authors.forename: Robert B
authors.initials: RB
authors.lastname: Belshe
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000097:blood*; D000914:Antibodies, Viral / Q000097:blood*; D002460:Cell Line; D002522:Chlorocebus aethiops; D003165:Complement System Proteins / Q000737:chemistry; D003412:Cricetulus; D005260:Female; D017930:Genes, Reporter; D006168:Guinea Pigs; D006558:Herpes Genitalis / Q000175:diagnosis / Q000276:immunology* / Q000821:virology; D006561:Herpes Simplex / Q000175:diagnosis / Q000276:immunology* / Q000821:virology; D018259:Herpesvirus 1, Human / Q000254:growth & development / Q000276:immunology*; D018258:Herpesvirus 2, Human / Q000254:growth & development / Q000276:immunology*; D057166:High-Throughput Screening Assays; D006801:Humans; D007106:Immune Sera / Q000737:chemistry; D007763:Lac Operon; D009500:Neutralization Tests; D011401:Promoter Regions, Genetic; D014709:Vero Cells; D014771:Virion / Q000276:immunology; D001616:beta-Galactosidase / Q000235:genetics / Q000276:immunology
chemical_list: D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D007106:Immune Sera; D003165:Complement System Proteins; D001616:beta-Galactosidase