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Journal of Biomolecular Screening
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High Throughput Fluorescence Polarization: A Homogeneous Alternative to Radioligand Binding for Cell Surface Receptors

Michael Allen

Receptor & Enzyme Screening Technologies, Glaxo Wellcome Medicines Research Centre, Stevenage, Herts, UK

Julian Reeves

Receptor Pharmacology, Glaxo Wellcome Medicines Research Centre, Stevenage, Herts, UK

Geoffrey Mellor

Molecular Discovery, Glaxo Wellcome Medicines Research Centre, Stevenage, Herts, UK

High throughput fluorescence polarization (FP) assays are described that offer a nonradioactive, homogeneous, and low-cost alternative to radioligand binding assays for cell surface receptors (G protein-coupled receptors and ligand-gated ion channels). FP assays were shown to work across a range of both peptide (vasopressin V1a and {delta}-opioid) and nonpeptide (ß-adrenoceptor, 5-hydroxytryptamine3) receptors. Structure-activity relationships were investigated at ß1-receptors and were found to be consistent with radioligand binding assays. FP was shown to tolerate up to 5% DMSO with no loss in sensitivity or signal window. From a random set of 1,280 compounds, 1.9% were found to significantly interfere with FP measurement. If fluorescent or quenching compounds were eliminated (3% of all compounds), less than 0.4% of compounds were found to interfere with FP measurement. Assays could be run in 384-well plates with little loss of signal window or sensitivity compared to 96-well plate assays. New advances in FP measurement have therefore enabled FP to offer a high throughput alternative to radioligand binding for cell surface receptors.

Journal of Biomolecular Screening, Vol. 5, No. 2, 63-69 (2000)
DOI: 10.1177/108705710000500202
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