Description
Interferometric Scattering (iSCAT) microscopy is a highly sensitive technique that measures the linear scattering signals of individual nanoparticles through image-based interferometric detection. However, the application of iSCAT to 3D particle tracking has been limited by the oscillation of the signal-to-noise ratio (SNR) when particles move along the axial direction. In this work, we introduce a strategy to overcome this limitation by evenly distributing the phase of a particle's scattered field using a spiral phase mask at the back pupil plane. Our approach, termed “spiral phase iSCAT microscopy (SP-iSCAT),” maintains a consistent SNR as particles move, thus enhancing the accuracy of particle localization in 3D. We evaluate the performance of SP-iSCAT through numerical simulations, benchmarking the theoretical limits. Additionally, we experimentally demonstrate high-precision, ultrahigh-speed 3D tracking of freely diffusing nanoparticles in water. We successfully measure the diffusion trajectories of particles as small as 20 nm in diameter at a high speed of 20,000 frames per second. The capability of accurate tracking of small particles by SP-iSCAT allows for precise quantification of hydrodynamic particle sizes at the single-particle level. Furthermore, SP-iSCAT provides quantitative measurements of the amplitude of the scattered signal, enabling the determination of particle polarizability. This combination of information allows for the direct assessment of particle size and mass density of individual nanoparticles in solution, opening the door to the investigation of biological nanoparticles in complex systems, such as cell vesicles and virus particles.
