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A cellular screening assay using analysis of metal-modified fluorescence lifetime

Cade, Nicholas I.; Fruhwirth, Gilbert; Archibald, Stephen J.; Ng, Tony; Richards, David

Authors

Nicholas I. Cade

Gilbert Fruhwirth

Tony Ng

David Richards

Abstract

Current methods for screening cell receptor internalization often require complex image analysis with limited sensitivity. Here we describe a novel bioassay based on detection of changes in global fluorescence lifetime above a gold substrate, with superresolution axial sensitivity and no need for image analysis. We show that the lifetime of enhanced green fluorescent protein expressed in a cellular membrane is greatly reduced in close proximity to the gold, resulting in a distance-dependent lifetime distribution throughout the cell. We demonstrate the application of this phenomenon in a screening assay by comparing the efficacies of two small molecule inhibitors interfering with the internalization process of a G protein-coupled receptor.

Journal Article Type Article
Publication Date Jun 2, 2010
Journal BIOPHYSICAL JOURNAL
Print ISSN 0006-3495
Electronic ISSN 1542-0086
Publisher Biophysical Society
Peer Reviewed Peer Reviewed
Volume 98
Issue 11
Pages 2752-2757
Institution Citation Cade, N. I., Fruhwirth, G., Archibald, S. J., Ng, T., & Richards, D. (2010). A cellular screening assay using analysis of metal-modified fluorescence lifetime. Biophysical journal, 98(11), 2752-2757. https://doi.org/10.1016/j.bpj.2010.03.016
DOI https://doi.org/10.1016/j.bpj.2010.03.016
Keywords CXCR4 antagonist, chemokine receptor, medicinal inorganic, cellular assay, FRET, fluorescence microscopy