| Solution Information | help | |
| Enzyme: | NADPH oxidase 1 | |
| inhibitor: | BDBM65841 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Assay Overview: The purpose of this cell-based assay is to determine the potency of compounds identified as active in the "Late stage results from the probe development effort to identify inhibitors of (NADPH oxidase 1) NOX1: HEK/293 Percent inhibition" (AID 1792) assay. This chemiluminescence assay employs a luminol probe to monitor intracellular ROS in HEK/293 cells. Protocol Summary: In this assay, HEK/293 cells seeded into 6-well plates were cotransfected with pRK5-Myc-Nox1, -NoxO1, -NoxA1, and pRK5-myc-Rac1CAQ61L. After 16 hours, test or control compounds were added, followed 2 hours later by luminal and horseradish peroxidase. The interaction of luminol with NOX1-generated ROS/superoxide inside cells yields an unstable endoperoxide that generates light, leading to increased well luminescence. As designed, compounds that inhibit cellular NOX1 activity will reduce intracellular ROS and endoperoxide levels, leading to reduced luminol-ROS interactions, reduced endoperoxide produc | |
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