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Jurkat cells were seeded on the same day at 300,000 cells/100uL/well in a Costar black wall/clear bottom 96-well plate. The ROS assay loading solution (100 uL/well) was added and incubated in a 5% CO2, 37°C incubator for 1 hour. And then the cells were treated with 1mM, 0.1mM H2O2 or without H2O2 for 30 minutes. The fluorescence signal was monitored at Ex/Em = 490/525 nm (cutoff at 515 nm) with bottom read mode using FlexStation (Molecular Devices).