Fluorescence Spectroscopy

Book chapter, 2016

Fluorescence spectroscopy is unique in its combination of sensitivity with experimental versatility. The advent of relatively affordable, robust, yet sophisticated, benchtop fluorimeters in conjunction with the vast and growing number of commercially available fluorescent probes have contributed to the accessibility and popularity of fluorescence spectroscopy. It has become one of the most important analytical techniques for the in vitro study of biomolecules and in vivo cellular imaging, providing spatial and temporal information. This chapter discusses the majority of techniques commonly used in the study of biomolecules. The quintessential fluorescence-based technique is steady-state fluorescence spectroscopy. In contrast to steady-state fluorescence spectroscopy, time-resolved fluorescence analysis can facilitate the simultaneous analysis of multiple emissive states with overlapping spectral bands, each with its own fluorescence decay, by deconvolution of a sample's multiexponential decay curve. Steady-state fluorescence spectroscopy, time-resolved fluorescence spectroscopy, and fluorescence anisotropy are typically concerned with monitoring a single fluorescent probe.