Differential Fluorescence Scanning Assay (DSF Assay) v1

Differential scanning fluorimetry (DSF) is a thermal shift assay technique used to measure the denaturation of proteins caused by increasing temperature, which breaks the non-covalent bonds that underlie protein folding. Meanwhile, the fluorescent dye, which gives high fluorescence in a nonpolar environment, is used in the DSF assay to probe the hydrophobic sites, which are generally embedded in the interior part of the proteins and exposed when proteins are denatured by high temperature. Thus, as the temperature increases, the ratio of denatured proteins increases, thereby elevating the intensity of the fluorescence. When the protein is fully unfolded, the intensity would peak. It can be used to test if the ligand-protein interaction occurs by detecting the shift of thermostability of the target protein after the addition of the small molecule ligand.