Fluorescent Labeling of Proteins

Many single-molecule experimental techniques exploit fluorescence as a tool to investigate conformational dynamics and molecular interactions or track the movement of proteins in order to gain insight into their biological functions. A prerequisite to these experimental approaches is to graft one or more fluorophores on the protein of interest with the desired photophysical properties. Here, we present detailed procedures for the current most efficient methods used to covalently attach fluorophores to proteins. Alternative direct and indirect labeling strategies are also described.