The FLUOR DE LYS® HDAC fluorometric activity assay kit is a sensitive and convenient alternative to protocols utilizing radiolabeled, acetylated histones or peptide/HPLC methods for the assay of histone deacetylases. It is based on the unique FLUOR DE LYS® (Fluorimetric Histone deAcetylaseLysyl) substrate and developer combination and provides an assay that can be carried out in two simple mixing steps, all on the same 96-well plate. Histone deacetylase inhibitors have shown promise as anti-tumor agents and naturally this has stimulated interest in the screening of compounds for HDAC inhibition.
- Useful for assaying lysates, immunoprecipitates or inhibitor screening using the nuclear extract provided
- HeLa nuclear extract, a rich source of HDACs 1 and 2 for use as a positive control or as a source of HDAC activity for screening
- Includes enough reagent for 100-200 assays
- Compatible with class I & IIb HDAC and sirtuins (with addition of NAD+)
The Fluor 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 Fluor de Lys® developer generates a fluorophore. The assay has been used successfully with preparations of all the known class I HDACs-HDAC1, HDAC2, HDAC3 and HDAC8 with class II HDACs 4-7, 9 and 10 and with the human Sir2 homolog, SIRT1. It has shown that the Fluor de Lys® substrate is cell-permeable and is deacetylated in situ by cellular HDACs. The deacetylated substrate accumulates inside cells and may be quantified by addition of Fluor de Lys® developer to a cell lysate.
Delivery information: Kit includes Nuclear Extract from HeLa Cells (human cerival cancer cell line), FLUOR DE LYS® Substrate (50 µl), 20X FLUOR DE LYS® Developer Concentrate (300 µl), Trichostatin A (HDAC Inhibitor) (100 µl), FLUOR DE LYS® Deacetylated Standard (30 µl), HDAC Assay Buffer (20 ml), ½ volume microplate, ½ volume white microplate.
