Particle Tracking Single Protein-Functionalized Quantum Dot Diffusion and Binding at Silica Surfaces

We evaluate commercial QD585 and QD605 streptavidin-functionalized quantum dots (QDs) for single-particle tracking microscopy at surfaces using total internal reflectance fluorescence and measure single QD diffusion and nonspecific binding at silica surfaces in static and flow conditions. The QD diffusion coefficient on smooth, near-ideal, highly hydroxylated silica surfaces is near bulk-solution diffusivity, as expected for repulsive surfaces, but many QD trajectories on rougher, less-than-ideal surfaces or regions display transient adsorptions. We attribute the binding to defect sites or adsorbates, possibly in conjunction with protein conformation changes, and estimate binding energies from the transient adsorption lifetimes. We also assess QD parameters relevant to tracking, including hydrodynamic radius, charge state, signal levels, blinking reduction with reducing solutions, and photoinduced blueing and bleaching.