| Assay Method Information | |
| | Scintillation Proximity Assay |
| Description: | The inhibition of a microsomal preparation of 11beta -HSD1 by compounds of the invention was measured essentially as previously described (K. Solly, S. S. Mundt, H. J. Zokian, G. J. Ding, A. Hermanowski-Vosatka, B. Strulovici, and W. Zheng, High-Throughput Screening of 11-Beta-Hydroxyseroid Dehydrogenase Type 1 in Scintillation Proximity Assay Format. Assay Drug Dev Technol 3 (2005) 377-384). All reactions were carried out at rt in 96 well clear flexible PET Microbeta plates (PerkinElmer). The assay begins by dispensing 49 ul of substrate solution (50 mM HEPES, pH 7.4, 100 mM KCl, 5 mM NaCl, 2 mM MgCl2, 2 mM NADPH and 160 nM [3H]cortisone (1 Ci/mmol)) and mixing in 1 uL of the test compounds in DMSO previously diluted in half-log increments (8 points) starting at 0.1 mM. After a 10 minute pre-incubation, 50 uL of enzyme solution containing microsomes isolated from CHO cells overexpressing human 11beta -HSD1 (10-20 ug/ml of total protein) was added. |
| Affinity data for this assay | |
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