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| | Receptor Calcium FLIPR Antagonist Assay |
| Description: | The assay measures intracellular changes of Ca2+ mediated by the synthetic probing agonist 3-{[2-(2-Trifluoromethyl-biphenyl-4-yl)-benzo[b]thiophen-5-ylmethyl]-amino}-propionic acid (GNF-AC-1) in the HeLa-S1P1/Galpha16 cell clone 1: HeLa (human cervix carcinoma, ATCC CCL2) cells stably expressing N-terminally myc-tagged human S1P1 receptors (GenBank accession No. NM001400; UNIPROT P21453) and promiscuous Galpha16 protein (GenBank accession number M63904, Swissprot P30679) are cultured at 37 C., 5% CO2, and 95% relative humidity. The cells are plated in 384 well black plates (10'000 cells per well). After 24 hours the cells are loaded with Fluo-4-AM (1.6 uM in HBSS and 2.5 mM probenicid) for 1 hour at 37 C. After washing, the cells are transferred to the FLIPR. The test compounds are added at different concentrations (<=100 uM) in HBSS in the presence of 0.1% BSA and changes in fluorescence are recorded (indication of agonism). |
| Affinity data for this assay | |
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