Time-Resolved Fluorescence

Fluorimetry has experienced an explosive growth since the early 1980s, much of which has been driven by the use of fluorescence as a noninvasive technique for biology and biochemistry. Fluorescence techniques are widely used to quantify molecular parameters of different chemical, biochemical, and biological processes because of their inherent sensitivity, specificity and temporal resolution. In fact, the luminescence lifetime is an important characteristic of a fluorescent molecule and its environment. Many intra- and intermolecular processes are able to modulate the molecule emission which cannot be investigated by steady-state fluorescence measurements. For example, rotational diffusion, resonance-energy transfer, or dynamic quenching occur on the same timescale as the fluorescence decay. Lifetime measurements reveal dynamic information on the nanosecond timescale that is useful in fundamental studies of quenching and energy transfer and can be used in analytical methodologies to enhance selectivity and/or to perform multicomponent determinations. For example, fluorescence quenching may occur due to a ground state reaction or an excited state reaction. Only by measuring the fluorescence lifetime may one determine which process causes the quenching.