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High-content screening of GPCR activation with MetaXpress, AcuityXpress and the Transfluor assay system

METAXPRESS APPLICATION NOTE

By Susan Catalano, Ph.D., Acumen Pharmaceuticals, Sylvia de Bruin, Joe Bosworth, Ph.D., Neal Gliksman, Ph.D., Kimberly Best, Ph.D. and Paula Rickert, Ph.D., Molecular Devices Corporation, 1311 Orleans Drive, Sunnyvale, CA 94089.


INTRODUCTION
G-protein coupled receptors (GPCRs are the largest class of pharmaceutical targets and, as a result, assays for detecting GPCR receptor agonists and antagonists play a major role in screening operations. Recently Xsira Pharmaceuticals introduced the Transfluor assay, a novel high-content assay that monitors GPCR activation by measuring GPCR desensitization and recycling. The Transfluor assay is now exclusively available from Molecular Devices.

The Transfluor Assay, validated on over 90 different GPCRs, uses a common pathway independent of downstream G-protein signaling which is shared by virtually all GPCRs, representing all classes of GPCRs and G proteins. The assay uses image analysis to quantitate internalization of GFP-tagged beta-arrestin that associates with the receptor of interest following activation.1 (See Figure 1.)


HIGH-CONTENT SCREENING OF GPCR ACTIVATION WITH METAXPRESS, ACUITYXPRESS AND THE TRANSFLUOR ASSAY SYSTEM

Unstimulated cells display diffuse cytoplasmic beta-arrestin-GFP fluorescence. When an over-expressed GPCR is activated, beta-arrestin targets the receptor for internalization, resulting in the appearance of small fluorescent clathrin-coated pits. If the receptor has a high affinity for beta-arresti
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