PI/RNase Staining Buffer

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	- Cells are fixed with 70% cold ethanol, and stored at −20 °C for 1 h - Cells were washed with PBS containing 0.1% triton X-100 on a shaking water bath at 37 °C, 40 rpm for 30 min.	- Store tubes at 4°C protected from light prior to analysing. Analyze on the flow cytometer within 1 hour.

Protocol tips
- Cells are fixed with 70% cold ethanol, and stored at −20 °C for 1 h - Cells were washed with PBS containing 0.1% triton X-100 on a shaking water bath at 37 °C, 40 rpm for 30 min.
Downstream tips
- Store tubes at 4°C protected from light prior to analysing. Analyze on the flow cytometer within 1 hour.

Publication protocol

Cell cycle analyses were performed after culturing A549 cells for 24 h. The cells were harvested, fixed with 70% cold ethanol, and stored at −20 °C for 1 h. They were washed twice with PBS and incubated with phosphate–citrate buffer and 0.1% triton X-100 on a shaking water bath (37 °C, 40 rpm) for 30 min. The cells were washed again with PBS prior to staining with 0.5 mL of PI/RNase staining buffer (BD Biosciences) for 15 min at room temperature. The cells were analyzed through flow cytometry. Cell cycle distribution was calculated from 10,000 cells by using ModFit LT software (Topsham, ME, USA)

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