| Solution Information | help | |
| Enzyme: | NADPH oxidase 1 | |
| inhibitor: | BDBM65842 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Assay Overview: The purpose of this cell-free assay is to evaluate the ability of compounds to inhibit ROS production by another cellular source, xanthine oxidase. This chemiluminescence assay employs a luminol probe to monitor ROS production. Protocol Summary: In this assay, xanthine oxidase (0.5 mg/mL) was dispensed to all wells of a 96-well plate followed by test or control compounds. Hypoxanthine was dispensed to all wells, followed by the addition of horseradish peroxide and luminal and reading luminescence. The interaction of luminol with xanthine oxidase-generated ROS/superoxide inside cells yields an unstable endoperoxide that generates light, leading to increased well luminescence. As designed, compounds that inhibit cellular xanthine oxidase activity will reduce intracellular ROS and endoperoxide levels, leading to reduced luminol-ROS interactions, reduced endoperoxide production, reduced light emission, and reduced well luminescence. Test compounds were assayed in tripli | |
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