Cell Research: |
Proc Natl Acad Sci U S A,2008 Feb 26;105(8):3041-6. | Discovery of a selective inhibitor of oncogenic B-Raf kinase with potent antimelanoma activity.[Pubmed: 18287029] |
Cell lines:COLO205, A375, WM2664, COLO829, HT716, SW620, H460, Calu-6, HCT116, SK-MEL2, SK-MEL3, Lovo, H1299, 1205Lu, and C8161 cells
Concentrations: Dissolved in DMSO, final concentrations ~1 mM
Incubation Time: 24, 48, and 72 hoursBR/>
Method:
Cells are treated with various concentrations PLX-4720 for 24, 48, and 72 hours. Cell proliferation is measured by using the CellTiter-Glo Luminescent Cell Viability Assay or MTT assay. For cell cycle analysis, supernatant and cells are collected, pelleted, and fixed with 70% ethanol. Before staining with propidium iodide (10 μg/mL), cells are incubated for 1 hour at 37 °C in 0.5 mg/mL RNase I to rid samples of residual RNA contamination. Samples are then analyzed by using the EPICS XL apparatus. For the assessment of apoptosis, media and cells are harvested and pelleted before staining with annexin-FITC and propidium iodide. Samples are subsequently analyzed by using the EPICS XL apparatus.
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