ICT’s Intracellular Total ROS Activity Assay provides a good screening option for assessing the potency of oxidative stress inhibitor and activator reagents, and will help to determine how oxidative stress modulates varied intracellular pathways. This product assesses the overall level of intracellular ROS activity and can be analyzed using a flow cytometer.
- Prepare samples and controls in 1X Assay Buffer or buffer of choice (0.5 mL at a cell density of 5 x 105 – 1 x 106 cells/mL).
- Reconstitute Total ROS Green with 100 µL DMSO to produce a 500X stock concentrate.
- Dilute 1:10 by adding 900 µL 1X Assay Buffer to the 500X stock Total ROS Green to produce a 50X working solution.
- Add Total ROS Green to each sample at 1:50. For example, add 10 µL into 490 µL cultured cells.
- Incubate 1 hour at 37°C.
- Treat cells with test compounds for desired period of time. Keep cells protected from light.
- Analyze with a flow cytometer. Total ROS Green excites at 490 nm and emits at 520 nm.
Kaproń B, Czarnomysy R, Wysokiński M, Andrys R, Musilek K, Angeli A, Supuran CT, Plech T. 1,2,4-Triazole-based anticonvulsant agents with additional ROS scavenging activity are effective in a model of pharmacoresistant epilepsy. J Enzyme Inhib Med Chem. 2020 Dec;35(1):993-1002. doi: 10.1080/14756366.2020.1748026. Full Text