| Assay Method Information | |
| | In Vitro Assay |
| Description: | An in vitro assay showed inhibition of CXCR2-mediated intracellular calcium release by treatment with compound 9 (IC50=586±91 nM). Briefly, human neutrophils were suspended in HBSS− (without Ca2+ and Mg2+) containing 10 mM HEPES and FLIPR Calcium 3 dye (3.1×107 cells in total volume 1.7 mL). Cells were aliquoted (200 μL of the cell suspension per tube, 8 tubes total) and 2 μL of compound 9 (with appropriate dilutions) were added to each of 6 tubes. The tested concentrations of compound 9 were 156 nM, 312 nM, 625 nM, 1250 nM, 2500 nM and 5000 nM. As controls, 2 μL of DMSO (1% final concentration) were added to 2 other tubes. Cells were incubated for 30 min at 37° C. After dye loading, tubes were centrifuged at 6,000 rpm for 1 min, supernatant was removed and the cell pellet was re-suspended in 200 μL of HBSS+ (with Ca2+ and Mg2+) containing 10 mM HEPES. |
| Affinity data for this assay | |
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