Continuous Monitoring Biosensing Mediated by Single-Molecule Plasmon-Enhanced Fluorescence in Complex Matrices

Continuous detection of critical markers directly at the point of interest and in undiluted biological fluids represents the next fundamental step in biosensing. The goal of realizing such a platform is utterly challenging because it requires a reversible biosensor that enables the tracking of pico- to nanomolar molecular concentrations over long time spans in a compact device. Here we describe a sensing method based on plasmon-enhanced fluorescence capable of single-molecule detection of unlabeled analyte by employing biofunctionalized gold nanoparticles. The very strong plasmon-enhanced fluorescence signals allow for single-molecule sensing in unaltered biological media, while the use of low-affinity interactions ensures the continuous tracking of increasing and decreasing analyte concentrations with picomolar sensitivity. We demonstrate the use of a sandwich assay for a DNA cancer marker with a limit of detection of picomolar and a time response of 10 min. The enhanced single-molecule signals will allow for miniaturization into a small and cheap platform with multiplexing capability for application in point-of-care diagnostics, monitoring of industrial processes, and safe keeping of the environment.