| Solution Information | help | |
| Enzyme: | Bifunctional epoxide hydrolase 2 | |
| inhibitor: | BDBM41938 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | sEH assay Materials 1. recombinant sEH, PHOME (substrate) and sEH inhibitors were provided by Dr. Bruce Hammock, University of California, Davis, CA. 2. Assay buffer: 25 mM Bis-Tris, pH 7.0, 0.1 mg/ml BSA 3. sEH working solution: 24 nM in assay buffer. 4. PHOME working solution: 58 uM (21.6 ug/ml), 1.2% DMSO in assay buffer. 5. Inhibitor working solution: 1.2 M ZnSO4 sEH screen protocol: 1. 10 ul of 24 nM sEH in assay buffer was manually dispensed into columns 1-12 of a black, flat-bottomed 96-well plate (Corning 3915) 2. 10 ul of the following solutions were dispensed: a. 16 % DMSO in Assay buffer to column 1 (no inhibitor control) b. 1.6 uM compound in 16% DMSO, in assay buffer into columns 2 # 11. c. 1.2 M ZnSO4 into column 12 (Inhibitor control). 3. Shake plate 20 sec to mix. 4. Incubate plate 10 min at room temperature. 5. 60 uL of PHOME working solutions was added to every well. 6. Shake plate 20 sec to mix. 7. Final concentrations of the components in the assay were as f | |
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