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- Simple two-step protocol with < 1 hour time to answer
- Colorimetric readout at 405 nm compatible with most plate readers
- Resistant to detergent interference common to antibody-based assays
- Eliminates need for radioactivity, extractions, and/or chromatography
- Suitable for high throughput analysis
Useful for assaying lysates, immunoprecipitates or inhibitor screening using the nuclear extract provided.- Includes HeLa nuclear extract, a rich source of HDACs 1 & 2 for use as a positive control or as a source of HDAC activity for screening.
Compatible with class I & IIb HDAC and sirtuins (with addition of NAD+).
The COLOR DE LYS® system (Colorimetric Histone de Acetylase Lysyl Substrate/Developer) is a sensitive and convenient alternative to protocols utilizing radiolabeled, acetylated histones or peptide/HPLC methods for the assay of histone deacetylases. The assay procedure has two steps. First, the COLOR DE LYS® 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, mixing with the COLOR DE LYS® developer causes an increase in yellow color intensity, and absorption at 405 nm.
Note: The COLOR DE LYS® substrate is efficiently deacetylated by HDAC1 and HDAC2, the major contributors to HDAC activity in HeLa nuclear extracts. It is however, a poor substrate for HDAC3 and recombinant human HDAC8 (BIOMOL International, unpublished results). The activities of other HDAC isotypes with the COLOR DE LYS® substrate have yet to be investigated. HDAC3 and HDAC8 do deacetylate the FLUOR DE LYS® substrate, which is the basis for the HDAC fluorometric activity assay kit (Prod. No. BML-AK500).
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Our Color-de-Lys HDAC colorimetric assay is less sensitive to detergents than competitive antibody-based assays. HeLa nuclear extract (8.3µg) was added to the substrate and buffer recommended by the manufacturer in the presence or absence of 0.1% Triton X-100. After 60 minutes at 37°C, the reaction was stopped and processed as recommended by the manufacturers. Triton X-100 showed little or no effect on the Color-de-Lys reaction, but caused an apparent 70% inhibition of the antibody-based assay.
Our COLOR DE LYS® HDAC colorimetric assay is less sensitive to detergents than competitive antibody-based assays. HeLa nuclear extract (8.3µg) was added to the substrate and buffer recommended by the manufacturer in the presence or absence of 0.1% Triton X-100. After 60 minutes at 37°C, the reaction was stopped and processed as recommended by the manufacturers. Triton X-100 showed little or no effect on the COLOR DE LYS® reaction, but caused an apparent 70% inhibition of the antibody-based assay.
Reaction Scheme of the HDAC Colorimetric Activity Assay (patent pending). Deacetylation of the substrate sensitizes it to the developer, which causes an increase in yellow intensity and absorption at 405 nm.
| Regulatory Status |
RUO – Research Use Only |
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Related Products
FLUOR DE LYS® HDAC fluorometric activity assay kit
BML-AK500
First-to-market by the leader in Epigenetics research tools
| Application | Activity assay, Fluorescent detection, HTS |
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