Sirt3 Assay Kit

The Fluorogenic SIRT3 Assay Kit is a complete assay system designed to measure Sirtuin 3 (SIRT3) activity for screening and profiling applications. It comes in a convenient 96-well format, with all the reagents necessary for 32 fluorescent SIRT3 activity measurements*. The kit includes purified SIRT3 enzyme and a SIRT inhibitor, Nicotinamide, for use as a positive and negative control, respectively. The Fluorogenic SIRT3 Assay Kit is based on a unique fluorogenic substrate and developer combination. This assay method eliminates dealing with the radioactivity, extraction, and chromatography aspects of traditional assays. Using this kit, only two simple steps on a microtiter plate are needed to analyze the SIRT3 activity level. First, the HDAC fluorometric substrate (HDAC substrate 1), containing an acetylated lysine side chain, is incubated with a sample containing SIRT3 activity (e.g. HeLa nuclear extract or purified SIRT3 enzyme). The deacetylation sensitizes the substrate so subsequent treatment with the Lysine Developer produces a fluorophore that can then be measured using a fluorescence reader. *Note: The kit includes sufficient HDAC substrate 1, buffers, NAD, nicotinamide, and detection reagents for a full 96-well plate. Researchers can test their own SIRT3-containing samples in the remaining wells, or additional SIRT3 enzyme may be ordered separately (Cat. #50014). Components: SIRT3 human recombinant enzyme, 70 µg, Fluorogenic SIRT substrate 1, 50 µl @ 5 mM, Nicotinamide Adenine Dinucleotide (NAD), 50 µl @ 50 mM, Nicotinamide, 500 µl @ 10 mM, 2x SIRT Developer (contains 2 mM Nicotinamide), 6 ml, SIRT assay buffer, 10 ml, 1 black, low binding NUNC black microtiter plate. References: 1. A. Ito et al. (2001) EMBO J. 20 1331. 2. N.A. Barlev et al. (2001) Mol. Cell 8 1243. 3. A. Ito et al. (2002) EMBO J. 21 6236.