ROS-ID® Total ROS/Superoxide detection kit

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	- After cells were treated with soluble RANKL for 6h, they are washed and cultured in a medium with 2% FBS. Cells were treated with oxidative stress detection reagent for 30 min on ice

Protocol tips
- After cells were treated with soluble RANKL for 6h, they are washed and cultured in a medium with 2% FBS. Cells were treated with oxidative stress detection reagent for 30 min on ice

Publication protocol

Intracellular ROS levels were detected using the Total ROS/Superoxide Detection Kit (Enzo Life Sciences Inc., Farmingdale, NY), according to the manufacturer's instructions. Cells were stimulated with soluble RANKL (sRANKL; Wako Pure Chemical Industries, Ltd.) for 6 h, washed, and collected in PBS supplemented with 2% fetal bovine serum (FBS). The cell suspension was incubated with oxidative stress detection reagent for 30 min on ice. After washing with PBS, intracellular ROS were detected using an Accuri C6 flow cytometer (BD Biosciences). The viable monocyte/macrophage fraction was gated on an FSC/SSC plot, and ROS levels were monitored in the FL-1 channel.

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