FRET enhancement in aluminum zero-mode waveguides

Area of Science:

• Biophysics
• Spectroscopy
• Nanotechnology

Background:

• Zero-mode waveguides (ZMWs) confine light to attoliter volumes, enabling single-molecule fluorescence studies at high concentrations.
• Förster resonance energy transfer (FRET) is a key technique in life sciences for studying molecular interactions.
• ZMWs offer potential for enhanced FRET measurements, but their influence on the FRET process requires careful evaluation.

Purpose of the Study:

• To quantify Förster resonance energy transfer (FRET) rates and efficiencies in individual donor-acceptor fluorophore pairs diffusing within aluminum zero-mode waveguides (ZMWs).
• To compare the effects of aluminum ZMWs versus gold ZMWs on FRET parameters.
• To understand how ZMWs influence FRET efficiency for single-molecule applications.

Main Methods:

• Diffusion of donor-acceptor fluorophore pairs through aluminum and gold zero-mode waveguides.
• Measurement of Förster resonance energy transfer (FRET) rates and efficiencies using fluorescence spectroscopy.
• Analysis of FRET rate dependence on donor decay rate and local density of optical states.

Main Results:

• FRET rates in both aluminum and gold ZMWs scale linearly with the isolated donor decay rate and the local density of optical states.
• Aluminum ZMWs provide greater fluorescence brightness enhancement factors compared to gold ZMWs due to lower optical losses in the green spectral region.
• Gold ZMWs exhibit a stronger influence on fluorescence decay rates.

Conclusions:

• The influence of zero-mode waveguides (ZMWs) on Förster resonance energy transfer (FRET) is quantifiable and predictable.
• Aluminum ZMWs are suitable for enhanced single-molecule FRET at physiological concentrations, offering a balance of brightness enhancement and minimal perturbation.
• Understanding ZMW-FRET interactions is crucial for developing advanced single-molecule analytical devices.