title: High-throughput ribozyme-based assays for detection of viral nucleic acids.
abstract: Many reports have suggested that target-activated ribozymes hold potential value as detection reagents. We show that a "half"-ribozyme ligase is activated similarly by three unstructured oligoribonucleotides representing the major sequence variants of a hepatitis C virus 5'-untranslated region (5'-UTR) target and by a structured RNA corresponding to the entire 5'-UTR. Half-ribozyme ligation product was detected both in an ELISA-like assay and in an optical immunoassay through the use of hapten-carrying substrate RNAs. Both assay formats afford a limit of detection of approximately 1 x 10(6) HCV molecules (1.6 attomol, 330 fM), a sensitivity which compares favorably to that provided by standard immunoassays. These data suggest that target-activated ribozyme systems are a viable approach for the sensitive detection of viral nucleic acids using high-throughput platforms.
authors.affiliation: Sirna Therapeutics Inc., 2950 Wilderness Place, Boulder, CO 80301 USA.
authors.forename: Karl
authors.initials: K
authors.lastname: Kossen
authors.forename: Narendra K
authors.initials: NK
authors.lastname: Vaish
authors.forename: Vasant R
authors.initials: VR
authors.lastname: Jadhav
authors.forename: Christopher
authors.initials: C
authors.lastname: Pasko
authors.forename: Hong
authors.initials: H
authors.lastname: Wang
authors.forename: Robert
authors.initials: R
authors.lastname: Jenison
authors.forename: James A
authors.initials: JA
authors.lastname: McSwiggen
authors.forename: Barry
authors.initials: B
authors.lastname: Polisky
authors.forename: Scott D
authors.initials: SD
authors.lastname: Seiwert
mesh_terms: D001483:Base Sequence; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006526:Hepatitis C / Q000175:diagnosis* / Q000821:virology; D007118:Immunoassay / Q000379:methods; D007700:Kinetics; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D016337:RNA, Catalytic / Q000378:metabolism*; D012367:RNA, Viral / Q000032:analysis*; D012680:Sensitivity and Specificity
chemical_list: D016337:RNA, Catalytic; D012367:RNA, Viral