title: Development and application of a flow cytometric potency assay for DNA vaccines.
abstract: We have developed a rapid, reliable, and sensitive quantitative flow cytometric assay to measure the in vitro potency and stability of DNA vaccines to be delivered either by particle-mediated epidermal delivery (PMED) or by electroporation. The method involves transfecting cells with test DNA and comparing the measured antigen expression to that generated with expression from known quantities of reference material DNA. The assay was adapted for performance under Good Laboratory Practice (GLP) guidelines and was successfully utilized to perform potency testing in support of a Phase I study for two hantavirus DNA vaccines delivered by gene gun. The results from the potency assays conducted over a 24-month period using this method proved to be highly reproducible with high signal-to-noise ratios. The assay was also adapted to assess the in vitro potency and stability of a DNA vaccine for Venezuelan equine encephalitis virus that will be delivered by electroporation. Our results indicate that this assay can be readily applied to support potency and stability testing of numerous DNA vaccines delivered by various methods, including multiagent vaccines.
authors.affiliation: U.S. Army Medical Research Institute of Infectious Diseases, FortDetrick, MD 21702, USA.
authors.forename: C V
authors.initials: CV
authors.lastname: Badger
authors.forename: J D
authors.initials: JD
authors.lastname: Richardson
authors.forename: R L
authors.initials: RL
authors.lastname: Dasilva
authors.forename: M J
authors.initials: MJ
authors.lastname: Richards
authors.forename: M D
authors.initials: MD
authors.lastname: Josleyn
authors.forename: L C
authors.initials: LC
authors.lastname: Dupuy
authors.forename: J W
authors.initials: JW
authors.lastname: Hooper
authors.forename: C S
authors.initials: CS
authors.lastname: Schmaljohn
mesh_terms: D000818:Animals; D000956:Antigens, Viral / Q000276:immunology; D019470:Biolistics; D019556:COS Cells; D002522:Chlorocebus aethiops; D016503:Drug Delivery Systems / Q000379:methods; D004355:Drug Stability; D004587:Electrophoresis, Agar Gel; D018274:Electroporation; D004666:Encephalitis Virus, Venezuelan Equine / Q000235:genetics / Q000276:immunology; D005434:Flow Cytometry / Q000295:instrumentation / Q000379:methods*; D006476:Orthohantavirus / Q000235:genetics / Q000276:immunology*; D006801:Humans; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D012015:Reference Standards; D015203:Reproducibility of Results; D014162:Transfection; D019444:Vaccines, DNA / Q000008:administration & dosage / Q000276:immunology*; D014765:Viral Vaccines / Q000008:administration & dosage / Q000276:immunology
chemical_list: D000956:Antigens, Viral; D019444:Vaccines, DNA; D014765:Viral Vaccines