Contributed by Daniel Daniel
Förster resonance energy transfer (FRET) is a mechanism that describes the transfer of energy between chromophores. A donor chromophore, which is in its electronically excited state, can transfer the energy non-radiatively to a acceptor chromophore that is nearby (typically less than 10 nm away) through dipole-dipole coupling. This process is known as FRET and it is exquisitely dependent on the distance between the two chromophores. FRET is often referred to as fluorescence resonance energy transfer, though this is a misnomer since the energy transfer is non-radiative and does not involve fluorescence. FRET allows imaging of objects that is separated by a distance of the order of 10 nm, which is way below the abbe diffraction limit of a microscope of 200 nm. It is one of the modern imaging techniques that allows us to surpass the diffraction limit that is characteristic of wide-field microscopy.
Figure 1. Schematic of FRET between the donor chromophore, cyan fluorescent protein (CFP), and the acceptor chromophore, yellow fluorescent protein (YFP). FRET will only occur when the two chromophores are sufficiently close to each other.
The principle of resonance energy transfer was first elucidated in the late 1940s by Theodor Förster. The idea was that if the fluorescent emission spectrum of the donor fluorophore overlaps with the absorption spectrum of the acceptor chromophore, and the two are near enough to each other, the donor fluorophore can transfer its excitation energy through long-range dipole-dipole interactions. This is a quantum mechanical process and does not require a collision and produces no heat. This is schematically shown in figure 2. Typically, the two chromophores have to be between 1-10 nm apart. At distance below 1 nm, other modes of electron and energy transfer becomes possible and beyond 10 nm, the probability of resonance energy transfer occuring becomes minimal.
In fact, the quantum yield of FRET process is given by
where <math>R_0</math> is the förster radius and the distance for which the efficiency falls to half. Typically, for fluorophores, <math>R_0</math> is of the order of 6 nm.
Figures 2 and 3 are taken from http://www.olympusfluoview.com/applications/fretintro.html