title: An automated time-of-drug-addition assay to routinely determine the mode of action of HIV-1 inhibitors.
abstract: Cell-based high-throughput screening campaigns are widely used to identify novel antiviral compounds, for example, against human immunodeficiency virus type 1 (HIV-1). Typically, these assays enable identification of compounds that potentially target any viral or cellular factor involved in the viral replication cycle. Unraveling the mechanism of action of these active compounds is an important step to facilitate further drug development. Time-of-addition (TOA) assays are an elegant tool to achieve this goal by comparing the TOA profile of novel compounds with those of well-studied reference compounds. Downscaling to a 384-well format and automation significantly increase the capacity of the TOA assay, enabling compound handling around the clock. Mechanical liquid dispensing with optimized time points for compound addition ensures robustness (Z'>0.8) and maximal resolution in profiling novel antiviral compounds. The presented methodology has been optimized for routine use and allows for fully automated high-throughput screening to support the process in search for novel inhibitors of HIV-1.
authors.affiliation: Janssen Infectious Diseases-Diagnostics BVBA , Beerse, Belgium .
authors.forename: Marnix
authors.initials: M
authors.lastname: Van Loock
authors.forename: Christel
authors.initials: C
authors.lastname: Van den Eynde
authors.forename: John
authors.initials: J
authors.lastname: Hansen
authors.forename: Peggy
authors.initials: P
authors.lastname: Geluykens
authors.forename: Tania
authors.initials: T
authors.lastname: Ivens
authors.forename: Sarah
authors.initials: S
authors.lastname: Sauviller
authors.forename: Lieve
authors.initials: L
authors.lastname: Bunkens
authors.forename: Koen
authors.initials: K
authors.lastname: Van Acker
authors.forename: Erik
authors.initials: E
authors.lastname: Nijs
authors.forename: Géry
authors.initials: G
authors.lastname: Dams
mesh_terms: D019380:Anti-HIV Agents / Q000008:administration & dosage / Q000494:pharmacology*; D001331:Automation; D001681:Biological Assay; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D003626:Data Display; D004305:Dose-Response Relationship, Drug; D015497:HIV-1 / Q000187:drug effects*; D057166:High-Throughput Screening Assays / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D007202:Indicators and Reagents; D012015:Reference Standards; D015203:Reproducibility of Results; D014779:Virus Replication / Q000187:drug effects
chemical_list: D019380:Anti-HIV Agents; D007202:Indicators and Reagents