[13] Fluorescence resonance energy transfer

Fluorescence resonance energy transfer (FRET) is a technique for measuring the distance between two points which are separated by approximately 10–75 A°. The technique is valuable because measurements can be made under physiological (or other) conditions with near angstrom resolution and with the exquisite sensitivity of fluorescence measurements. For these reasons, FRET has found wide use in polymer science, biochemistry, and structural biology. The extent of energy transfer can be measured because the fluorescence of the donor (both intensity and lifetime) decreases, or is quenched, and the acceptor, if fluorescent, increases its fluorescence, or becomes “sensitized,” with energy transfer. These changes in fluorescence can be measured by comparing a complex labeled with both donor and acceptor to ones labeled only with donor and only with acceptor. The ability to measure intensities and lifetimes of both donor and acceptor emission with high accuracy and excellent signal-to-background, coupled with the unusually large R0s, makes luminescence resonance energy transfer a potentially powerful technique for measuring distances in biological systems.