BD Cycletest™ Plus DNA Kit

Protocol tips

Upstream tips	Protocol tips	Downstream tips
- Cells were seeded and treated with 2,4′,6-trihydroxy-4-methoxybenzophenone (at IC50) at 37°C for 24 h, 48 h, and 72 h.	- Cells were fixed with ethanol, washed in warm PBS, and incubated at −20°C, overnight

Upstream tips
- Cells were seeded and treated with 2,4′,6-trihydroxy-4-methoxybenzophenone (at IC50) at 37°C for 24 h, 48 h, and 72 h.
Protocol tips
- Cells were fixed with ethanol, washed in warm PBS, and incubated at −20°C, overnight

Publication protocol

HT-29 cells were placed in Petri dishes at a density of 1 × 105 cells/well and grown for 24 h and then the cells were treated with 2,4′,6-trihydroxy-4-methoxybenzophenone (at IC50) at 37°C for 24 h, 48 h, and 72 h. After incubation, the cells were collected by trypsinization, fixed in ethanol, washed in warm PBS, and incubated at −20°C, overnight. Next, the cells were washed with 1 mL of cold PBS and stained with propidium iodide according to the Cycle TEST PLUS DNA Reagent Kit (Becton Dickinson, USA) protocol. The cells were sorted in a FACScalibur flow cytometer (BD, USA) using CellQuest Pro software (Becton Dickinson, USA) and a quantitative analysis of the cell cycle distribution was performed using a trial version of the ModFit LT software, version 4.0.

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