title: Automation of a phospho-STAT5 staining procedure for flow cytometry for application in drug discovery.
abstract: Drug discovery often requires the screening of compound libraries on tissue cultured cells. Some major targets in drug discovery belong to signal transduction pathways, and PerFix EXPOSE* allows easy flow cytometry phospho assays. We thus investigated the possibility to further simplify and automate this assay, to allow the direct screening of drugs targeting signaling pathways. We show here the sensitivity of this fully automated assay on human growth hormone (hGH)-driven JAK/STAT5-activated IM-9 cells, and we discuss the throughput of this system, which is compatible with medium-throughput drug screening. Because the kit works directly on whole blood samples, ex-vivo assays are also possible with this approach, which could allow for the screening of drugs under more physiological conditions.
authors.affiliation: Beckman Coulter Life Science, Inc., Global Assay and Applications Development, Marseille, France.
authors.forename: Fabrice
authors.initials: F
authors.lastname: Malergue
authors.affiliation: Beckman Coulter Life Science, Inc., Global Assay and Applications Development, Marseille, France.
authors.forename: Andreas
authors.initials: A
authors.lastname: van Agthoven
authors.affiliation: Beckman Coulter Life Science, Inc., Global Assay and Applications Development, Marseille, France.
authors.forename: Caroline
authors.initials: C
authors.lastname: Scifo
authors.affiliation: Department of Cell Biology, Center for Molecular Medicine, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Dave
authors.initials: D
authors.lastname: Egan
authors.affiliation: Department of Cell Biology, Center for Molecular Medicine, University Medical Center Utrecht, Utrecht, The Netherlands gstrous@umcutrecht.nl.
authors.forename: Ger J
authors.initials: GJ
authors.lastname: Strous
mesh_terms: D001331:Automation; D002460:Cell Line; D055808:Drug Discovery / Q000295:instrumentation / Q000379:methods*; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation / Q000379:methods; D005434:Flow Cytometry / Q000295:instrumentation / Q000379:methods*; D057166:High-Throughput Screening Assays / Q000295:instrumentation / Q000379:methods; D019382:Human Growth Hormone / Q000494:pharmacology; D006801:Humans; D053612:Janus Kinases / Q000378:metabolism; D010539:Permeability / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D015203:Reproducibility of Results; D050799:STAT5 Transcription Factor / Q000378:metabolism*; D054852:Small Molecule Libraries
chemical_list: D050799:STAT5 Transcription Factor; D054852:Small Molecule Libraries; D019382:Human Growth Hormone; D053612:Janus Kinases
keywords: GH-driven; JAK; STAT; cell-based; flow cytometry; high throughput; whole blood