Upgrading interferometric scattering microscopy with ensemble statistical analysis

Interferometric scattering (iSCAT) microscopy, label-free high-speed (up to ~1000 frames per second) imaging and tracking technique, has proven to be a versatile tool by measuring the mass and 3D position of nanoparticles and biomolecules as well as visualizing real-time dynamics of nanoscale events in complex cellular environments. However, the quantification of iSCAT signals has not been straightforwardly defined in practical terms. We delve into several issues associated with signal processing in iSCAT: error-prone post-processing routine and lack of statistical reliability in the convention of iSCAT contrast. After providing a brief account of concepts and principles of correlation spectroscopy, we here discuss an alternative ensemble (higher number density of scatterers) statistical analysis that can be used to extract the dynamic information of scattering particles from fluctuating iSCAT signals. Finally, our perspective on the correlation approach toward time-correlated iSCAT technique will be presented.