Extending Single-Molecule Förster Resonance Energy Transfer (FRET) Range beyond 10 Nanometers in Zero-Mode Waveguides
Förster resonance energy transfer (FRET) is widely used as a molecular ruler to monitor biomolecular conformations and interactions dynamics. However, an intrinsic limitation in FRET is that the signal decreases very rapidly with the dye to dye distance, and FRET is generally not detectable anymore beyond 10 nm.
In a recent paper published in ACS Nano, we use nanoapertures milled in an aluminum film (called zero-mode waveguides ZMW) to overcome the spatial range limit in FRET. Our optimized structures creates favorable conditions to enhance the FRET efﬁciency by 3-fold at a large donor-acceptor distance of 13.6 nm, well beyond the classical Förster radius.
- We demonstrate that ZMWs can extend the spatial range of FRET to distances where dipole-dipole interactions would otherwise be too weak to produce detectable FRET signals.
- ZMWs can be combined with molecular constructs featuring multiple acceptor dyes to further improve the FRET efficiency and extend the spatial range.
- General guidelines are discussed for performing quantitative FRET measurements inside ZMWs and nanoapertures and apply our approach for biophysics and biochemistry applications.
Also freely available on ArXiv 1907.03734