Pi rnase staining kits

HepG2 cells were plated in 6-well plates at a density of 1 × 10⁶ cells per well. After attachment, cells were treated with PP-26 for 24 hours. After treatment with PP-26, the HepG2 cells were centrifuged at 650 g for 10 minutes and resuspended in PBS, and then stained by PI/RNase staining kits (Multisciences, Hangzhou, China) or annexin V fluorescein solution (Beyotime, Shanghai, China) at room temperature for 10 minutes. To assess onset of apoptosis, propidium iodide in HEPES buffer was added. Cell cycle stage and onset of apoptosis was analyzed by flow cytometry (FCM, Becton Dickinson, Franklin Lakes, NJ, USA), in accordance with the manufacturer’s protocol. The percentages of cells in G0/G1, S, and G2/M phases, or the ratios of apoptotic cells, were calculated.

Cells were harvested and washed with ice-cold phosphate-buffered saline (PBS). The PI/RNase staining kits (Multi-sciences, Shanghai, China) and an annexin V-fluorescein isothiocyanate propidium iodide (FITC/PI) apoptosis detection kit (Multisciences, Shanghai, China) were used to detect cell cycle and apoptosis according to the manufacturer’s protocol using a FACScan instrument (BD, Franklin Lakes, NJ, USA), respectively. Three independent experiments were performed.