Interferometric Phase Intensity Nanoscopy (iPINE), Revealing Nanostructural Features, Resolves Proximal Nanometer Objects below the Diffraction Limit: Implications in Long-Time Super-Resolution of Biological Dynamics

Abstract

The ability to spatially and temporally map nanoscale environments in situ over extended time scales would be transformative for biology, biomedicine, and bioengineering. All nanometer objects, from nanoparticles down to single proteins, scatter light. Interferometric scattering stands as a powerful tool, offering ultrasensitivity and resolution vital for visualizing nanoscale entities. Interferometric scattering from an individual nanoparticle down to an individual protein has been detected; however, resolving adjacent nanometer objects with interferometric scattering has not yet been demonstrated. In this work, we present interferometric phase intensity nanoscopy (iPINE) to resolve adjacent nanometer objects with interferometric scattering. We demonstrate that multiphase and sensitivity of iPINE reveal ellipse Airy patterns correlated with nanostructural features. We show that eliminating background fluctuation by employing circularly polarized illumination in iPINE is essential to separate proximal nanometer objects below the diffraction limit. We envision iPINE for resolving a variety of adjacent nanometer objects from nanoparticles down to proximal proteins in situ over extended time periods for a wide range of applications in biology, biomedicine, and bioengineering. We expect iPINE to be especially important in applications of biological dynamics that require extended observation times.

Two-color interferometric scattering (iSCAT) microscopy reveals structural dynamics in discrete plasmonic molecules

“…iSCAT in combination with a phase-sensitive detection has also shown promise to resolve sub-diffraction limit contacts between NPs with separations down to about 150 nm. 36 All of these properties make iSCAT a promising strategy to investigate the assembly of sub-40 nm NP into plasmonic molecules and monitor separations within these assemblies on chemical length scales.…”