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Apoptosis analysis with Annexin V/PI staining
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Apoptotic cells were detected using Annexin V-FITC Apoptosis Detection Kit (Thermo Fisher Scientific) as instructed by the manufacturer. Briefly, cells were seeded in 24-well plates and cultured for approximately 24 h to reach a confluence of nearly 80%. Cells were then stained with the Annexin V-FITC and PI solution for 15 min, and analyzed using the FACSCalibur to determine the percentage of apoptotic cells, which were represented as the percentage sum of early apoptotic (Annexin V+PI−) and late apoptotic (Annexin V+PI+) cells.
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