| Assay Method Information | |
| | Enzymatic Assays |
| Description: | Histone deactylase 3 (HDAC3):The HDAC3 assay kit measures HDAC activity using a fluorogenic substrate containing an acetylated lysine. In a 96-well plate, substrate (final concentration 20 mM) was mixed with bovine serum albumin (1 mg/mL) and assay buffer. A 10× solution of the test inhibitor was added, keeping a final 1% DMSO concentration. The reaction was initiated with the addition of HDAC3 enzyme (final concentration 2 ng/well), and the plate was incubated at 37° C. for 30 min. Controls consist of substrate control (no enzyme added), enzyme control (assay buffer with DMSO vehicle), and inhibitor control (final concentration of 2 mM trichostatin A). HDAC developer solution was then added to each well and incubated at room temperature for 15 min. Solution fluorescence was read at 360ex/450em nm on a plate reader. Assay was performed in triplicate, and IC50 values were determined from a graph of percent inhibition as a function of concentration using GraphPad. |
| Affinity data for this assay | |
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