| Assay Method Information | |
| | Measurement of 11beta-HSD1 Activity |
| Description: | The enzyme assay was performed in a round-bottom 96-well plate. The enzyme was pre-incubated in the assay buffer in the presence of NADPH and inhibitor. Next, the reaction is initiated by adding a regenerating system (consisting of glucose-6-phosphate, glucose-6-phosphate dehydrogenase, and MgCl2), and labeled 3H-cortisone substrate. After the incubation, the substrate and product peaks are separated and analyzed by using an HPLC system. The initial reaction velocities recorded were in the linear range and were determined by measuring the peak area for cortisol formation with time. The inhibition of 11beta-HSD was analyzed by fitting the data to an equation to determine apparent inhibition constant Ki. |
| Affinity data for this assay | |
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