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Assay Method Information

Assay Name:  Inhibition Assay
Description:  This example demonstrates reversible inhibition of USP1/UAF1 by an embodiment of the invention.The compound (81) at a concentration of ten times its 1050 value was pre-incubated with 10 μM USP1/UAF1 at room temperature for 15 min. This solution was then diluted by 100-fold in assay buffer containing 50 mM HEPES, pH 7.8, 0.1 mg/mL BSA, 0.5 mM EDTA, and 1 mM DTT and incubated at room temperature for an additional 15 min. Next, 20 μM K63-linked diubiquitin was added into the solution and incubated at 37 C. for 10 min. Laemmli sample buffer was added to quench the reaction. The reaction products were then separated on a 20% denaturing SDS-PAGE gel and stained with Coomassie Blue. USP1/UAF1 incubated with DMSO was treated as 100% USP1/UAF1 activity.
Affinity data for this assay
 

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Last update November 1, 2007
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