Human SIRT1 is the homolog of yeast Sir2 and has been shown to regulate the activity of the p53 tumor suppressor and inhibit apoptosis. Cayman’s SIRT1 FRET-based Screening Assay provides a convenient fluorescence-based method for screening SIRT1 inhibitors or activators. The procedure requires only two easy steps, both performed in the same microplate. In the first step, the substrate, which is coupled to the fluorophore and quencher, is incubated with human recombinant SIRT1 along with its cosubstrate NAD+. Deacetylation sensitizes the substrate such that treatment with the developer in the second step results in the separation of the quencher and fluorophore. The resulting fluorescence is analyzed using an excitation wavelength between 335-345 nm and emission wavelength between 440-465 nm.