title: High-throughput screening with HyperCyt flow cytometry to detect small molecule formylpeptide receptor ligands.
abstract: High-throughput flow cytometry (HTFC), enabled by faster automated sample processing, represents a promising high- content approach for compound library screening. HyperCyt is a recently developed automated HTFC analysis system by which cell samples are rapidly aspirated from microplate wells and delivered to the flow cytometer. The formylpeptide receptor (FPR) family of G protein-coupled receptors contributes to the localization and activation of tissue-damaging leukocytes at sites of chronic inflammation. Here, the authors describe development and application of an HTFC screening approach to detect potential anti-inflammatory compounds that block ligand binding to FPR. Using a homogeneous no-wash assay, samples were routinely processed at 1.5 s/well (approximately 2500 cells analyzed/sample), allowing a 96-well plate to be processed in less than 2.5 min. Assay sensitivity and accuracy were validated by detection of a previously documented active compound with relatively low FPR affinity (sulfinpyrazone, inhibition constant [K(i)]=14 microM) from among a collection of 880 compounds in the Prestwick Chemical Library. The HyperCyt system was therefore demonstrated to be a robust, sensitive, and highly quantitative method with which to screen lead compound libraries in a 96-well format.
authors.affiliation: Cytometry and Department of Pathology, Cancer Research and Treatment Center, University of New Mexico, Health Sciences Center, Albuquerque, NM 87131, USA.
authors.forename: Susan M
authors.initials: SM
authors.lastname: Young
authors.forename: Cristian
authors.initials: C
authors.lastname: Bologa
authors.forename: Eric R
authors.initials: ER
authors.lastname: Prossnitz
authors.forename: Tudor I
authors.initials: TI
authors.lastname: Oprea
authors.forename: Larry A
authors.initials: LA
authors.lastname: Sklar
authors.forename: Bruce S
authors.initials: BS
authors.lastname: Edwards
mesh_terms: D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D008024:Ligands; D044042:Receptors, Formyl Peptide / Q000378:metabolism*; D012680:Sensitivity and Specificity; D020298:U937 Cells
chemical_list: D008024:Ligands; D044042:Receptors, Formyl Peptide