Characterizing GPCR Activation Using Automated Live Cell Imaging
Poster Jul 24, 2017
Joe Clayton and Peter Banks
G protein coupled receptor (GPCR)-mediated pathways are critical for cells to respond to intercellular and environmental cues, and are a major focus of drug discovery efforts, particularly for cancer treatment. The molecules that activate GPCRs, and the resulting signaling cascades triggered by associated G proteins, are diverse. Fluorescent dyes and biosensors can be used to monitor changes in second messenger levels, including Ca2+ and cyclic AMP (cAMP), in response to GPCR activation. Here we describe a live cell imaging based approach to detect GPCR activation using the Lionheart™ FX Automated Live Cell Imager and Gen5™ Microplate Reader and Imager Software. This method provides a large assay window and improved sensitivity over methods relying on total fluorescence intensity measurements. Dual in-line dispense tips enable addition of GPCR agonists with continuous monitoring of cellular response. Additionally, an image capture rate of up 20 frames per second enables characterization of rapid GPCR kinetics.
Despite the developments in conventional PCR, the complexity of multiplex Real Time PCR is still limited due to the lack of sufficient detection channels. To achieve high-end multiplexing capacity on standard Real Time PCR machines, Anapa Biotech has developed the MeltPlex® technology (see box on right).READ MORE
Genome-wide association studies (GWAS) have identified more than 100 genetic loci associated with type 2 diabetes. The majority of these are located in the intergenic or intragenic regions suggesting that the implicated variants may alter chromatin conformation. This, in turn, is likely to influence the expression of nearby or more remotely located genes to alter beta cell function. At present, however, detailed molecular and functional analyses are still lacking for most of these variants. We recently analysed one of these loci and mapped five causal variants in an islet-specific enhancer cluster within the STARD10 gene locus. Here, we aimed to understand how these causal variants influence b-cell function by alteration of the chromatin structure of enhancer clusterREAD MORE