Photobleaching

Photo - a chemical change in the fluorophore molecule that permanently limits the ability to fluoresce. The covalent bonds or binding interactions between the fluorophore and the molecule (carrier) are broken by the energy from the incident photons. This phenomenon becomes a problem when live samples are repeatedly imaged. Reduction of bleaching is achieved by using highly sensitive cameras (e.g. EM CCD) and low excitation radiation intensities. There are various methods that exploit this phenomenon to study the motion and diffusion of molecules - FRAP, FLIP.