Interference scattering (iSCAT) microscopy of nanoobjects

Abstract

Label-free analysis made possible through the advent of scattering-based imaging techniques offers a viable alternative to fluorimetry. While dark field microscopy collects pure scattered light, common-path approaches like interferometric scattering microscopy (iSCAT) rely on augmentation of the weak scattering signal with a strong reference wave. These approaches allow for super-resolution, single-molecule investigation of molecular properties and behaviors with high precision and accuracy. The project includes the construction of an iSCAT microscope and subsequent ultrasensitive imaging and characterization of individual nanoparticles as well as proteins as proof of concept. This thesis focuses on characterizing and optimizing technical parameters in the experimental setup as well as combining iSCAT microscopy with a photothermal method for imaging of static plasmonic particles. Moreover, various approaches for signal enhancement are theoretically modelled comprising multiple factors connected to the recording device, objective and nanopositioning stage. In order to characterize the setup, exemplary results from dynamical iSCAT measurements of 10 nm gold nanoparticles are presented. Finally, the iSCAT setup was expanded to include a heating beam for plasmonic excitation of gold nanoparticles (GNPs) embedded in a thermo-optical matrix. Photothermal images of GNPs down to a size of 20 nm were reconstructed and analyzed at different modulation frequencies.