title: Highly sensitive multiplex assay for detection of human immunodeficiency virus type 1 and hepatitis C virus RNA.
abstract: Various nucleic acid assays have been developed and implemented for diagnostics and therapeutic monitoring of human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) infections. The high-throughput, semiautomated assays described here were developed to provide a method suitable for screening plasma specimens for the presence of HIV-1 and HCV RNAs. Three assays were developed: a multiplex HIV-1/HCV assay for simultaneous detection of HIV-1 and HCV, and discriminatory assays for specific detection of HIV-1 and HCV. The assay systems utilize three proprietary technologies: (i) target capture-based sample preparation, (ii) transcription-mediated amplification (TMA), and (iii) hybridization protection assay (HPA). An internal control is incorporated into each reaction to control for every step of the assay and identify random false-negative reactions. The assays demonstrated a sensitivity of at least 100 copies/ml for each target, and they detected with similar sensitivity all major variants of HCV and HIV-1, including HIV-1 group O strains. Assay sensitivity for one virus was not affected by the presence of the other. The specificity of these TMA-driven assays was >or=99.5% in both normal donor specimens and plasma containing potentially interfering substances or other blood-borne pathogens. Statistical receiver operating characteristic plots of 1 - specificity versus sensitivity data determined very wide analyte cutoff values for each assay at the point at which the assay specificity and sensitivity were both >or=99.5%. The sensitivity, specificity, and throughput capability predict that these assays will be valuable for large-volume plasma screening, either in a blood bank setting or in other diagnostic applications.
authors.affiliation: Research and Development, Gen-Probe Incorporated, San Diego, California 92121, USA. cristig@gen-probe.com
authors.forename: C
authors.initials: C
authors.lastname: Giachetti
authors.forename: J M
authors.initials: JM
authors.lastname: Linnen
authors.forename: D P
authors.initials: DP
authors.lastname: Kolk
authors.forename: J
authors.initials: J
authors.lastname: Dockter
authors.forename: K
authors.initials: K
authors.lastname: Gillotte-Taylor
authors.forename: M
authors.initials: M
authors.lastname: Park
authors.forename: M
authors.initials: M
authors.lastname: Ho-Sing-Loy
authors.forename: M K
authors.initials: MK
authors.lastname: McCormick
authors.forename: L T
authors.initials: LT
authors.lastname: Mimms
authors.forename: S H
authors.initials: SH
authors.lastname: McDonough
mesh_terms: D001782:Blood Donors; D015658:HIV Infections / Q000150:complications / Q000175:diagnosis / Q000821:virology; D015497:HIV-1 / Q000235:genetics* / Q000302:isolation & purification*; D016174:Hepacivirus / Q000235:genetics* / Q000302:isolation & purification*; D006526:Hepatitis C / Q000150:complications / Q000175:diagnosis / Q000821:virology; D006801:Humans; D008403:Mass Screening; D012367:RNA, Viral / Q000032:analysis* / Q000235:genetics*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014773:Virology / Q000379:methods* / Q000706:statistics & numerical data
chemical_list: D012367:RNA, Viral