Photobleaching is the photochemical destruction of a fluorophore. In microscopy, photobleaching may complicate the observation of fluorescent molecules, since they will eventually be destroyed by the light exposure necessary to stimulate them into fluorescing. This is especially problematic in time-lapse microscopy. A fluorophore is a component of a molecule which causes a molecule to be fluorescent. ... It has been suggested that this article or section be merged into microscope. ...

However, photobleaching may also be exploited to study the motion and/or diffusion of molecules, for example via the FRAP technique. Principle of FRAP Fluorescence recovery after photobleaching (FRAP) is a technique used in cellular imaging where a fluorochrome attached to a molecule is destroyed on purpose with an intense flash of light (by a laser). ...

Loss of activity caused by photobleaching can be controlled by reducing the intensity or time-span of light exposure, by increasing the concentration of fluorophores, or by employing more robust fluorophores that are less prone to bleaching. To a reasonable approximation, a given molecule will be destroyed after a constant exposure (intensity of emission X emission time X number of cycles) because, in a constant environment, each absorption-emission cycle has an equal probability of causing photobleaching.

lifetime

Depending on the material, dyes can produce different photon numbers and therefore have different lifetimes (at e.g. 10⁵ photons/s):

• Green fluorescent protein: 10⁴-10⁵; 0.1-1 s
• Typical organic dye: 10⁵-10⁶; 1-10 s
• CdSe/ZnS Quantum dot: 10⁸; > 1000 minutes

GFP ribbon diagram from PDB database The green fluorescent protein (GFP) is a protein from the jellyfish Aequorea victoria that fluoresces green when exposed to blue light. ... Fluorescence induced by exposure to ultraviolet light in vials containing various sized Cadmium selenide (CdSe) quantum dots. ...

External links

Introduction to Optical Microscopy an article about photobleaching