The HDAC Colorimetric Assay/Drug Discovery Kit is a complete assay system designed to measure histone deacetylase (HDAC) activity in cell or nuclear extracts, immunoprecipitates or purified enzymes. It comes in a convenient 96-well format, with all reagents necessary for Colorimetric HDAC activity measurements and calibration of the assay. In addition, a HeLa nuclear extract, rich in HDAC activity, is included with the kit. The extract is useful as either a positive control or as the source of HDAC activity for inhibitor/drug screening. Also included is the potent HDAC inhibitor, Trichostatin A, which may be used as model inhibitor.
The HDAC Colorimetric Assay/Drug Discovery Kit is based on the unique Substrate and Developer combination. The system (Colorimetric Histone deAcetylase Lysyl Substrate/Developer) is a highly sensitive and convenient alternative to radiolabeled, acetylated histones or peptide/HPLC methods for the assay of histone deacetylases. The assay procedure has two steps. First, the Substrate, which comprises an acetylated lysine side chain, is incubated with a sample containing HDAC activity (HeLa nuclear or other extract, purified enzyme, bead-bound immunocomplex, etc.). Deacetylation of the substrate sensitizes the substrate so that, in the second step, treatment with the Developer causes an increase in yellow intensity and absorption at 405nm. There is a linear correlation between the absorption and the amount of dye released within instrument limitations.