The ultimate detection limit in analytic chemistry and biology is the single molecule. Commonly, fluorescent dye labels or enzymatic amplification are employed. This requires additional labeling of the analyte, which modifies the species under investigation and therefore influences biological processes. Here, we utilize single gold nanoparticles to detect single unlabeled proteins with extremely high temporal resolution. This allows for monitoring the dynamic evolution of a single protein binding event on a millisecond time scale. The technique even resolves equilibrium coverage fluctuations, opening a window into Brownian dynamics of unlabeled macromolecules. Therefore, our method enables the study of protein folding dynamics, protein adsorption processes, and kinetics as well as nonequilibrium soft matter dynamics on the single molecule level.

Efficient and cost-effective multiplexed detection schemes for proteins in small liquid samples would bring drastic advances to fields like disease detection or water quality monitoring. We present a novel multiplexed sensor with randomly deposited aptamer functionalized gold nanorods. The spectral position of plasmon resonances of individual nanorods, monitored by dark-field spectroscopy, respond specifically to different proteins. We demonstrate nanomolar sensitivity, sensor recycling, and the potential to upscale to hundreds or thousands of targets.

We use plasmon rulers to follow the conformational dynamics of a single protein for up to 24 h at a video rate. The plasmon ruler consists of two gold nanospheres connected by a single protein linker. In our experiment, we follow the dynamics of the molecular chaperone heat shock protein 90 (Hsp90), which is known to show “open” and “closed” conformations. Our measurements confirm the previously known conformational dynamics with transition times in the second to minute time scale and reveals new dynamics on the time scale of minutes to hours. Plasmon rulers thus extend the observation bandwidth 3–4 orders of magnitude with respect to single-molecule fluorescence resonance energy transfer and enable the study of molecular dynamics with unprecedented precision.