title: Multiplexed reporter gene assays: monitoring the cell viability and the compound kinetics on luciferase activity.
abstract: High-throughput screening assays with multiple readouts enable one to monitor multiple assay parameters. By capturing as much information about the underlying biology as possible, the detection of true actives can be improved. This report describes an extension to standard luciferase reporter gene assays that enables multiple parameters to be monitored from each sample. The report describes multiplexing luciferase assays with an orthogonal readout monitoring cell viability using reduction of resazurin. In addition, this technical note shows that by using the luciferin substrate in live cells, an assay time course can be recorded. This enables the identification of nonactive or unspecific compounds that act by inhibiting luciferase, as well as compounds altering gene expression or cell growth.
authors.affiliation: Novartis Institute for Biomedical Research, Basel, Switzerland. marie-cecile.didiot@novartis.com
authors.forename: Marie-Cecile
authors.initials: MC
authors.lastname: Didiot
authors.forename: Sergio
authors.initials: S
authors.lastname: Serafini
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Pfeifer
authors.forename: Frederick J
authors.initials: FJ
authors.lastname: King
authors.forename: Christian N
authors.initials: CN
authors.lastname: Parker
mesh_terms: D000890:Anti-Infective Agents / Q000494:pharmacology; D001548:Benzalkonium Compounds / Q000494:pharmacology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D003513:Cycloheximide / Q000494:pharmacology; D049454:Firefly Luciferin / Q000032:analysis; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D007700:Kinetics; D008156:Luciferases / Q000235:genetics* / Q000378:metabolism*; D010078:Oxazines / Q000378:metabolism; D014966:Xanthenes / Q000378:metabolism
chemical_list: D000890:Anti-Infective Agents; D001548:Benzalkonium Compounds; D010078:Oxazines; D014966:Xanthenes; C005843:resazurin; D049454:Firefly Luciferin; D003513:Cycloheximide; D008156:Luciferases