| Solution Information | help | |
| Enzyme: | Neuropeptides B/W receptor type 1 | |
| inhibitor: | BDBM75945 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Assay Overview: The purpose of this assay is to determine dose response curves for synthesized compounds in a 384-well plate format for antagonism of GPR7. Although GPR7 is naturally coupled to Gai, which decreases cAMP levels upon activation, this assay employs a chimeric cell line that forces the receptor to use Gqi3, and therefore the assay readout is calcium release. In this assay HEK cells stably co-transfected with the human GPR7 receptor and Gaqi3 (hGPR7 HEK293T/Gqi3 cell line) are treated with test compounds, followed by measurement of intracellular calcium as monitored by the FLUO-8 fluorescent, cell permeable calcium indicator dye. As designed, compounds that act as GPR7 antagonists will decrease calcium mobilization, resulting in decreased relative fluorescence of the indicator dye, and thus decreased well fluorescence. Test compounds were assayed in triplicate in an 8-point 1:3 dilution series starting at a nominal test concentration of 20 uM. Protocol Summary: The hGPR | |
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