title: Application of the trak-C HCV core assay for monitoring antiviral activity in HCV replication systems.
abstract: The Ortho trak-C immunoassay has recently established detection of the HCV core antigen as a viable indirect marker of HCV replication in clinical samples. In this study, trak-C is used to monitor HCV replication in three pre-clinical models: the cellular HCV replicon system, transient transfection of HCV genomes, and the murine Alb-uPa/SCID HCV infection model. All of these systems utilize full-length HCV genomes that direct the expression of core, facilitating its detection with monoclonal antibodies. When performed with purified protein, the assay detects HCV core with a lower limit of detection at 1.5pg, and exhibits linear detection up to 100pg. When assaying extracts prepared from Huh-7 clone 21-5 cells harboring a full-length HCV replicon, core is detectable from as few as 63 cell equivalents. The assay was used to determine the sensitivity of Huh 21-5 cells to the antiviral effects of interferon (IFN). Inhibition by IFN-alpha using core detection was comparable to that observed using branched-DNA (bDNA 3.0) detection of HCV RNA. Replication of transfected full-length HCV 1a Con1 genomes in Huh-7 cells was also detectable using the trak-C assay. Finally, in the transgenic murine HCV infection model, the course of viral amplification was detected from serum using trak-C with kinetics similar to those observed with RNA detection. Given its ease of use and the lack of requirement for RNA purification, the trak-C assay has several advantages over RNA-based methods of viral monitoring.
authors.affiliation: Johnson and Johnson Pharmaceutical Research and Development, L.L.C., 3210 Merryfield Row, San Diego, CA 92121, USA.
authors.forename: Laurence
authors.initials: L
authors.lastname: Cagnon
authors.forename: Pamela
authors.initials: P
authors.lastname: Wagaman
authors.forename: Ralf
authors.initials: R
authors.lastname: Bartenschlager
authors.forename: Thomas
authors.initials: T
authors.lastname: Pietschmann
authors.forename: Tiejun
authors.initials: T
authors.lastname: Gao
authors.forename: Norman M
authors.initials: NM
authors.lastname: Kneteman
authors.forename: David L J
authors.initials: DL
authors.lastname: Tyrrell
authors.forename: Chander
authors.initials: C
authors.lastname: Bahl
authors.forename: Patrick
authors.initials: P
authors.lastname: Niven
authors.forename: Stephen
authors.initials: S
authors.lastname: Lee
authors.forename: Kenneth A
authors.initials: KA
authors.lastname: Simmen
mesh_terms: D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D002999:Clone Cells; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000706:statistics & numerical data; D015870:Gene Expression / Q000187:drug effects; D016679:Genome, Viral; D016174:Hepacivirus / Q000187:drug effects* / Q000235:genetics / Q000502:physiology*; D006526:Hepatitis C / Q000188:drug therapy / Q000821:virology; D006801:Humans; D007370:Interferon Type I / Q000494:pharmacology; D051379:Mice; D016513:Mice, SCID; D008822:Mice, Transgenic; D011994:Recombinant Proteins; D012680:Sensitivity and Specificity; D014758:Viral Core Proteins / Q000032:analysis* / Q000235:genetics; D014766:Viremia / Q000188:drug therapy / Q000821:virology; D014773:Virology / Q000379:methods / Q000706:statistics & numerical data; D014779:Virus Replication / Q000187:drug effects*
chemical_list: D000998:Antiviral Agents; D007370:Interferon Type I; D011994:Recombinant Proteins; D014758:Viral Core Proteins; C069130:nucleocapsid protein, Hepatitis C virus