| Assay Method Information | |
| | FLIPR Calcium 6 Assay |
| Description: | The dye is diluted with assay buffer (20mM HEPES in 1x HBSS, PH7.4); Probenecid was added to the final concentration of 5 mM; Hi. Vortexed vigorously for 1-2 minutes. Medium is removed from cell plate by flicking the cell plate on towel papers. 10 pl of assay buffer and 10 pl of 2*dye solution is added to each well of the cell plate. The cell plate is placed on plate shaker, the plate was agitated at 600rpm for 2 minutes. The plate is incubated at 37°C for 2 hours followed by additional 15-minute incubation at 25°C. 3*compound in assay buffer is prepared: a. Reference compounds are diluted to required concentration with DMSO. The compounds were added to a 384-well compound plate; b. Serial dilutions are performed; c. 10mM test compounds are added to the compound plate, and 3-fold serial dilutions were performed, d. Transferee 60 nl/well of compounds from source plate to a 384-well compound plate (Coming, 3657) by using an Echo; e. Add 20pl/well assay buffer to the compound plate; f. Mix the plate on plate shaker for 2 mins; The cell plate, compound plate and tips are put into FLIPR, 10pl of 3x compound is transferred to the cell plate per well with FLIPR. |
| Affinity data for this assay | |
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