Interferometric scattering enables fluorescence-free electrokinetic trapping of single nanoparticles in free solution

Anti-Brownian traps confine single particles in free solution by closed-loop feedback forces that directly counteract Brownian motion. The extended-duration measurement of trapped objects allows detailed characterization of photophysical and transport properties, as well as observation of infrequent or rare dynamics. However, this approach has been generally limited to particles that can be tracked by fluorescent emission. Here we present the Interferometric Scattering Anti-Brownian ELectrokinetic trap (ISABEL trap), which uses interferometric scattering rather than fluorescence to monitor particle position. By decoupling the ability to track (and therefore trap) a particle from collection of its spectroscopic data, the ISABEL trap enables confinement and extended study of single particles that do not fluoresce, that only weakly fluoresce, or which exhibit intermittent fluorescence or photobleaching. This new technique significantly expands the range of nanoscale objects that may be investigated at the single-particle level in free solution.Comment: Manuscript and SI; videos available upon reques

Ratiometric Sensing of Redox Environments Inside Individual Carboxysomes Trapped in Solution

Diffusion of biological nanoparticles in solution impedes our ability to continuously monitor individual particles and measure their physical and chemical properties. To overcome this, we previously developed the interferometric scattering anti-Brownian electrokinetic (ISABEL) trap, which uses scattering to localize a particle and applies electrokinetic forces that counteract Brownian motion, thus enabling extended observation. Here we present an improved ISABEL trap that incorporates a near-infrared scatter illumination beam and rapidly interleaves 405 and 488 nm fluorescence excitation reporter beams. With the ISABEL trap, we monitored the internal redox environment of individual carboxysomes labeled with the ratiometric redox reporter roGFP2. Carboxysomes widely vary in scattering contrast (reporting on size) and redox-dependent ratiometric fluorescence. Furthermore, we used redox sensing to explore the chemical kinetics within intact carboxysomes, where bulk measurements may contain unwanted contributions from aggregates or interfering fluorescent proteins. Overall, we demonstrate the ISABEL trap's ability to sensitively monitor nanoscale biological objects, enabling new experiments on these systems