title: Application of Micro Arrayed Compound Screening (microARCS) to identify inhibitors of caspase-3.
abstract: Micro Arrayed Compound Screening (microARCS) is a miniaturized ultra-high-throughput screening platform developed at Abbott Laboratories. In this format, 8,640 discrete compounds are spotted and dried onto a polystyrene sheet, which has the same footprint as a 96-well plate. A homogeneous time-resolved fluorescence assay format (LANCE) was applied to identify the inhibitors of caspase-3 using a peptide substrate labeled with a fluorescent europium chelate and a dabcyl quencher. The caspase-3 enzyme was cast into a thin agarose gel, which was placed on a sheet containing test compounds. A second gel containing caspase substrate was then laid above the enzyme gel to initiate the reaction. Caspase-3 cleaves the substrate and separates the europium from the quencher, giving rise to a time-resolved fluorescent signal, which was detected using a ViewLux charge-coupled device imaging system. Potential inhibitors of caspase-3 appeared as dark spots on a bright fluorescent background. Results from the microARCS assay format were compared to those from a conventional 96-well plate-screening format.
authors.affiliation: Abbott Laboratories, Pharmaceutical Products Division, Abbott Park, IL 60064, USA. Sujatha.M.Gopalakrishnan@abbott.com
authors.forename: Sujatha M
authors.initials: SM
authors.lastname: Gopalakrishnan
authors.forename: Jarkko
authors.initials: J
authors.lastname: Karvinen
authors.forename: James L
authors.initials: JL
authors.lastname: Kofron
authors.forename: David J
authors.initials: DJ
authors.lastname: Burns
authors.forename: Usha
authors.initials: U
authors.lastname: Warrior
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000032:analysis / Q000378:metabolism; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D005063:Europium / Q000378:metabolism; D005453:Fluorescence; D006801:Humans; D008904:Miniaturization / Q000295:instrumentation / Q000379:methods
chemical_list: D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D005063:Europium; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases