Plasmon-assisted Förster resonance energy transfer at the single-molecule level in the moderate quenching regime.

Metallic nanoparticles were shown to affect Förster energy transfer between fluorophore pairs. However, to date, the net plasmonic effect on FRET is still under dispute, with experiments showing efficiency enhancement and reduction. This controversy is due to the challenges involved in the precise positioning of FRET pairs in the near field of a metallic nanostructure, as well as in the accurate characterization of the plasmonic impact on the FRET mechanism.

DNA Origami Nanoantennas with over 5000-fold Fluorescence Enhancement and Single-Molecule Detection at 25 μM.

Optical nanoantennas are known to focus freely propagating light and reversely to mediate the emission of a light source located at the nanoantenna hotspot. These effects were previously exploited for fluorescence enhancement and single-molecule detection at elevated concentrations. We present a new generation of self-assembled DNA origami based optical nanoantennas with improved robustness, reduced interparticle distance, and optimized quantum-yield improvement to achieve more than 5000-fold fluorescence enhancement and single-molecule detection at 25 μM background fluorophore concentration.

Quantum yield and excitation rate of single molecules close to metallic nanostructures.

The interaction of dyes and metallic nanostructures strongly affects the fluorescence and can lead to significant fluorescence enhancement at plasmonic hot spots, but also to quenching. Here we present a method to distinguish the individual contributions to the changes of the excitation, radiative and non-radiative rate and use this information to determine the quantum yields for single molecules. The method is validated by precisely placing single fluorescent dyes with respect to gold nanoparticles as well as with respect to the excitation polarization using DNA origami nanostructures.

DNA origami-based standards for quantitative fluorescence microscopy.

Validating and testing a fluorescence microscope or a microscopy method requires defined samples that can be used as standards. DNA origami is a new tool that provides a framework to place defined numbers of small molecules such as fluorescent dyes or proteins in a programmed geometry with nanometer precision. The flexibility and versatility in the design of DNA origami microscopy standards makes them ideally suited for the broad variety of emerging super-resolution microscopy methods.

Single-molecule positioning in zeromode waveguides by DNA origami nanoadapters.

Nanotechnology is challenged by the need to connect top-down produced nanostructures with the bottom-up world of chemistry. A nanobiotechnological prime example is the positioning of single polymerase molecules in small holes in metal films, so-called zeromode waveguides (ZMWs), which is required for single-molecule real-time DNA sequencing. In this work, we present nanoadapters made of DNA (DNA origami) that match the size of the holes so that exactly one nanoadapter fits in each hole.