ABSTRACT
Fluorescence microscopy has been a quintessential and enabling tool for life scientists for more than a century and, as such, a source of major discoveries in biology. A fluorescence molecule has to fulfill many requirements in order to be most useful for a particular in vivo study. Thus, it is helpful to understand the basic principles underlying fluorescence signal generation. The fluorescence lifetime, i.e. average duration a molecule spends in the excited state before it experiences radiative relaxation, is of critical importance because it defines the shortest time scale in which dynamic events can be imaged. The interest in endogenous fluorophores dates back to the early twentie century due to the observed correlation between autofluorescence signal changes and the progression of malignant diseases. Exogenous fluorophores are the workhorse of modern biology, as they enable one to visualize and monitor a wide range of inter- and intracellular activities, up to the domain of whole-body imaging of small animals and clinical studies.
