Polarization Structured Illumination Microscopy

Fluorescence polarization microscopy (FPM) images both the intensity and orientation of fluorescent dipoles, which play a vital role in studying the molecular structures and dynamics of biocomplexes. However, current techniques have difficulty resolving the dipole assemblies on subcellular structures and their dynamics at the super-resolution (SR) scale. Structured illumination microscopy (SIM) is usually favored for long-term SR imaging of live cells due to its high spatiotemporal resolution, moderate labeling requirements and low photon budget. Moreover, perfect linear polarization light is required in SIM to obtain a high-contrast illumination pattern, making the SIM system an inherent FPM. In this chapter, we will introduce polarized structured illumination microscopy (pSIM), which achieves SR imaging of dipoles by interpreting the dipoles in spatio-angular hyperspace. We demonstrate the application of pSIM on a series of biological filamentous systems, such as cytoskeleton networks, and report the dynamics of short actin sliding across a myosin-coated surface. Furthermore, pSIM reveals the side-by-side organization of the actin ring structures in the membrane-associated periodic skeleton of hippocampal neurons and records the dipole dynamics of microtubules in live U2OS cells. pSIM is compatible with existing commercial and home-built SIM systems with various imaging modes, without hardware modification requirements.