FLUORESCENCE LIFETIME BIOSENSING: ENTERING THE MAINSTREAM

Publisher Summary This chapter emphasizes the salient features of florescence lifetime biosensing. When a fluorophore (fluorescent molecule or atom) is raised to an excited state by absorbance of a photon, it spends a very short time (typically a few nanoseconds) in that state before emitting a photon (or decaying by another process). For an individual fluorophore the time interval between absorption and emission varies randomly, but for an ensemble of fluorophores the average amount of time spent in the excited state is a characteristic of the fluorophore called the lifetime, т. The measurement of fluorescence lifetime biosensing can be done using a time-domain lifetime measurement approach or a frequency-domain lifetime measurement approach. This chapter also discusses the basic approaches for lifetime-based biosensing. While time- and frequency-domain instruments have been available commercially, they were not operable by scientists outside the field without significant training. However, the availability of essentially turnkey instrumentation has brought lifetime-based sensing much more into the mainstream. Finally, adaptation of the approach for use with the microscope and with optical fibers makes these sensors uniquely useful for biology and environmental monitoring; of particular importance is their utility for quantitative fluorescence resonance energy transfer (FRET)–based studies in live cells.