Fluorescence Lifetime Imaging, a New Tool in Confocal Microscopy

Abstract

It was not until the 1930s that measurement of the fluorescence lifetime (τf) and the confirmation of theory on this phenomenon became possible (Pringsheim, 1949). Before that time only phosphorescence lifetimes had been measured, evidently because this phenomenon is one or more orders of magnitude slower. The distinction between fluorescence and phosphorescence was first made by the observation of the afterglow. Emission having a noticeable afterglow was called phosphorescence. All other processes that did not have a noticeable afterglow were called fluorescence. Later, the distinction between the two was based on quantum mechanics. Phosphorescence is the process going through the higher-energy metastable triplet state having a low probability for the transition to the ground state and thus is slower. The fluorescence phenomenon only involves singlet states with transition probabilities that are usually higher, making the process faster.