Forster-Resonance Energy Transfer between Diffusing Molecules and a Functionalized Plasmonic Nanopore

Plasmonic nanopores are the subject of extensive investigations as a potential platform to enable efficient optical readout in translocation experiments with biomolecules such as DNA and proteins. They allow for the engineering of electromagnetic fields at the nanoscale, which are typically used to enhance the emission efficiency of fluorescent molecules. Their features make them suitable for detection strategies based on the energy transfer between translocating molecules and the nanopore itself. Here, we carry out an optical experiment to show that a handful of diffusing dyes acting as donors can exchange energy via Forster-resonance energy transfer (FRET) with a gold nanopore functionalized with dyes behaving as acceptors. The FRET pair is composed of ATTORho6G (donor) and Alexa610 (acceptor). To perform this proof-of-concept experiment, we use a gold nanopore with a diameter of 80 nm, prepared on a Si3N4 membrane. We observe that the presence of the acceptors on the walls of the nanopore reduces the lifetime of the diffusing donors. In addition, we observe that the presence of the acceptors reduces the fluorescence signal on the donor detection channel and increases the fluorescence signal on the acceptor detection channel. The combination of these three effects gives us enough evidence to claim that the diffusing donors exchange energy with the functionalized nanopore via FRET, despite the relatively large size of the nanopore. The FRET efficiency of the process is found to be of the order of 30%, which is in fairly good agreement with a theoretical value obtained using a simplified model.