Assay Method Information

Assay Name:  Enzyme Inhibition Assay
Description:  A reaction was performed on a 384-well plate (Greiner Bio One) using a reaction volume of 24 µL, and all the samples were diluted with an assay buffer (50 mM tris buffer, pH 7.4, 10% glycerol). 0.8 mM NADPH, 6 mM glucose 6-phosphate, 0.35 units/mL glucose 6-phosphate dehydrogenase (Sigma Chemical), and 3 mM magnesium chloride and a microsome fraction as an enzyme source were added to the plate, and a solution containing a test compound dissolved in a dimethyl sulfoxide/methanol solution was added at a final concentration of 0.1%. After the addition of the test compound, cortisone at a final concentration of 160 nM was added to start the reaction, and the reaction was performed at room temperature for 3 h. 25 µL of 100 mM carbenoxolone (Sigma Chemical) was added to terminate the reaction, and the amount of cortisol produced was measured with RUBYstar (BMG LABTECH JAPAN Ltd.) using a cortisol prototype kit (Cisbio International) according to the package insert.
Affinity data for this assay
 

If you find an error in this entry please send us an E-mail