title: Development of an in vitro antigen-detection test as an alternative method to the in vivo plaque reduction neutralization test for the quality control of Japanese encephalitis virus vaccine.
abstract: Japanese encephalitis virus (JEV) causes diseases that attack the human central nervous system. Traditionally, the quality control for JEV vaccines, in which the plaque reduction neutralization (PRN) titer is measured by the national control laboratories before the vaccine batches are marketed, has required laboratory animal testing. However, classical animal tests have inherent problems, including the very fact that animals are used, ethical issues, and the possibility of error. In this study, JEV antigen was measured in an in vitro assay to assess the feasibility of replacing in vivo assays that measure the PRN titers of JEV vaccines. We constructed a double-sandwich enzyme-linked immunosorbent assay (DS-ELISA) that could detect JEV envelope (E). Initially, monoclonal antibodies (mAbs) directed against the JEV E protein were generated and characterized. We isolated 18 mAbs against JEV E protein, and most were the IgG1 or IgG2a isotype. The mAbs (5F15 and 7D71) were selected as the most suitable mAb pair to detect JEV E protein. DS-ELISA with this pair detected as little as approximately 3 μg/mL JEV E protein and demonstrated a relationship between the amount of JEV E protein and the PRN titer. From these results, we surmise that this DS-ELISA may be useful, not only in terms of measuring the amount of JEV E protein, but also as a substitute for the PRN test for JEV vaccine evaluation.
authors.affiliation: National Center for Lot Release, Korea Food & Drug Administration, Chungcheongbuk-do, Korea.
authors.forename: Do Keun
authors.initials: DK
authors.lastname: Kim
authors.forename: Hye-Youn
authors.initials: HY
authors.lastname: Kim
authors.forename: Joo-Young
authors.initials: JY
authors.lastname: Kim
authors.forename: Michael B
authors.initials: MB
authors.lastname: Ye
authors.forename: Kee-Bum
authors.initials: KB
authors.lastname: Park
authors.forename: Euiri
authors.initials: E
authors.lastname: Han
authors.forename: Jaeok
authors.initials: J
authors.lastname: Kim
authors.forename: Sang
authors.initials: S
authors.lastname: Ja Ban
authors.forename: Seung Hwa
authors.initials: SH
authors.lastname: Hong
authors.forename: Yong Keun
authors.initials: YK
authors.lastname: Park
authors.forename: Jae-Hwan
authors.initials: JH
authors.lastname: Nam
mesh_terms: D000818:Animals; D000956:Antigens, Viral / Q000032:analysis* / Q000276:immunology; D004664:Encephalitis Virus, Japanese / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification / Q000502:physiology; D004672:Encephalitis, Japanese / Q000276:immunology* / Q000517:prevention & control / Q000821:virology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D022321:Japanese Encephalitis Vaccines / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000592:standards*; D051379:Mice; D008807:Mice, Inbred BALB C; D009500:Neutralization Tests; D011786:Quality Control; D014759:Viral Envelope Proteins / Q000032:analysis / Q000276:immunology; D010948:Viral Plaque Assay
chemical_list: D000956:Antigens, Viral; D022321:Japanese Encephalitis Vaccines; D014759:Viral Envelope Proteins