title: Characterization and use of mammalian-expressed vaccinia virus extracellular membrane proteins for quantification of the humoral immune response to smallpox vaccines.
abstract: The licensed smallpox vaccine Dryvax is used as the standard in comparative immunogenicity and protection studies of new smallpox vaccine candidates. Although the correlates of protection against smallpox are unknown, recent studies have shown that a humoral response against the intracellular mature virion and extracellular enveloped virion (EV) forms of vaccinia virus is crucial for protection. Using a recombinant Semliki Forest virus (rSFV) vector system, we expressed a set of full-length EV proteins for the development of EV antigen-specific enzyme-linked immunosorbent assays (ELISAs) and the production of monospecific antisera. The EV-specific ELISAs were used to evaluate the EV humoral response elicited by Dryvax and the nonreplicating modified vaccinia virus Ankara (MVA) in mouse vaccination experiments comparing doses and routes of vaccination. Quantitatively similar titers of antibodies against EV antigens A33R, A56R, and B5R were measured in mice vaccinated with Dryvax and MVA when MVA was administered at a dose of 10(8) plaque-forming units. Further, a substantial increase in the EV-specific antibody response was induced in mice inoculated with MVA by using a prime-boost schedule. Finally, we investigated the abilities of the EV-expressing rSFV vectors to elicit the production of polyclonal monospecific antisera against the corresponding EV proteins in mice. The monospecific serum antibody levels against A33R, A56R, and B5R were measurably higher than the antibody levels induced by Dryvax. The resulting polyclonal antisera were used in Western blot analysis and immunofluorescence assays, indicating that rSFV particles are useful vectors for generating monospecific antisera.
authors.affiliation: Laboratory of DNA Viruses, Division of Viral Products, Center for Biologics and Evaluation and Research/FDA, 1401 Rockville Pike, HFM-457, Rockville, MD 20892, USA. alonzo.garcia@fda.hhs.gov
authors.forename: Alonzo D
authors.initials: AD
authors.lastname: García
authors.forename: Clement A
authors.initials: CA
authors.lastname: Meseda
authors.forename: Anne E
authors.initials: AE
authors.lastname: Mayer
authors.forename: Arunima
authors.initials: A
authors.lastname: Kumar
authors.forename: Michael
authors.initials: M
authors.lastname: Merchlinsky
authors.forename: Jerry P
authors.initials: JP
authors.lastname: Weir
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D002522:Chlorocebus aethiops; D006224:Cricetinae; D004797:Enzyme-Linked Immunosorbent Assay; D005822:Genetic Vectors; D006367:HeLa Cells; D006801:Humans; D007114:Immunization; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D011995:Recombination, Genetic; D012672:Semliki forest virus / Q000235:genetics* / Q000378:metabolism; D012900:Smallpox Vaccine / Q000276:immunology*; D014616:Vaccinia virus / Q000378:metabolism; D014709:Vero Cells; D014771:Virion / Q000276:immunology / Q000378:metabolism
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; C078786:DryVax vaccine; D008565:Membrane Proteins; D012900:Smallpox Vaccine