High Content Compatible Fast Frame Rate Imaging of GPCR mediated Calcium Signaling


S Letzsch1; A Foitzik1; J Trask1A Schreiner1
1 PerkinElmer, UK


G-protein coupled receptors (GPCRs) are the largest family of plasma membrane receptors. They are activated by a huge variety of different molecules outside the cell and activate intracellular signal transduction pathways, which ultimately lead to different cellular responses, such as the increase of intracellular calcium concentration. Dysregulation of GPCR signaling, particularly fluctuations in calcium homeostasis signaling, has been implicated in different diseases, e.g. cancer and Alzheimer’s disease. Currently about 34% of all FDA approved drugs target GPCRs indicating their importance for therapeutic treatments.-. Thus, in vitro cellular GPCR measurements are considered a hallmark assay to understand the kinetics and mechanisms of actions to identify new drugs. Here we show how to analyze fast changes in intracellular calcium levels upon GPCR stimulation and inhibition using a fluorescence imaging assay on the Opera Phenix® Plus high-content screening system. Compounds are added using the on-board liquid handling module which transfers an agonist solution from a compound plate to the assay plate. Acquisition frame rates of up to 100 frames per second allow the monitoring of rapid changes in intracellular calcium concentration and dedicated image analysis tools of Harmony® software enable quantification of information from these image sequences

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