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Assay Method Information

Assay Name:  Biochemical Assay
Description:  The autoparsylation activity of the TNKS 1/2 or PARP1/2 enzymes was measured by the liquid chromatography-mass spectrometry (LC/MS) detection of nicotinamide as readout. Compound activity in inhibiting the TNKS and PARP autoparsylation was evaluated by IC50 measurements. In the compound screening assays, the reaction is composed of 5 μL of compound in 8-point serial dilutions with concentrations ranging from 0.0086 to 18.75 μM, 20 nM of purified enzyme, and 250 μM of β-NAD+ in the 1 Assay Buffer. After 60 min incubation at room temperature, the reactions were quenched by the addition of 10 μL of 5 quenching solution (20% formic acid and 500 nM [d]-nicotinamide in water). For the background control wells, 10 μL of the 5 quenching solution per well was added prior to the addition of β-NAD+.
Affinity data for this assay
 

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Last update November 1, 2007
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