Quenching (fluorescence)
In
Quenching is made use of in
Mechanisms
Förster resonance energy transfer
There are a few distinct mechanisms by which energy can be transferred non-radiatively (without absorption or emission of photons) between two dyes, a donor and an acceptor. Förster resonance energy transfer (FRET or FET) is a dynamic quenching mechanism because energy transfer occurs while the donor is in the excited state. FRET is based on classical dipole-dipole interactions between the transition dipoles of the donor and acceptor and is extremely dependent on the donor-acceptor distance, R, falling off at a rate of 1/R6. FRET also depends on the donor-acceptor spectral overlap (see figure) and the relative orientation of the donor and acceptor transition dipole moments. FRET can typically occur over distances up to 100 Å.
Dexter electron transfer
Dexter (also known as Dexter exchange or collisional energy transfer, colloquially known as Dexter Energy Transfer) is another dynamic quenching mechanism.[12] Dexter electron transfer is a short-range phenomenon that falls off exponentially with distance (proportional to e−kR where k is a constant that depends on the inverse of the van der Waals radius of the atom[citation needed]) and depends on spatial overlap of donor and quencher molecular orbitals. In most donor-fluorophore–quencher-acceptor situations, the Förster mechanism is more important than the Dexter mechanism. With both Förster and Dexter energy transfer, the shapes of the absorption and fluorescence spectra of the dyes are unchanged.
Dexter electron transfer can be significant between the dye and the solvent especially when hydrogen bonds are formed between them.
Exciplex
Static quenching
The remaining energy transfer mechanism is static quenching (also referred to as contact quenching). Static quenching can be a dominant mechanism for some reporter-quencher probes. Unlike dynamic quenching, static quenching occurs when the molecules form a complex in the ground state, i.e. before excitation occurs. The complex has its own unique properties, such as being nonfluorescent and having a unique
Collisional quenching
Collisional quenching occurs when the excited fluorophore experiences contact with an atom or molecule that can facilitate non-radiative transitions to the ground state. ... Excited-state molecule collides with quencher molecule and returns to ground state non-radiatively.
See also
- Dark quencher, for use in molecular biology.
- Förster resonance energy transfer, a phenomenon on which some quenching techniques rely