FIP Virtual Seminar "Multidimensional & Multifunctional Super-Resolution Microscopy"

Recent advances in super-resolution fluorescence microscopy have led to exciting spatial resolutions of ~10 nm in the far field. We are advancing beyond the structural (shape) information of super-resolution methods, to reveal (intracellular) functional parameters, including chemical polarity, diffusivity, and reactivity, with nanoscale resolution and single-molecule sensitivity. With spectrally resolved single-molecule localization microscopy, we encode functional parameters into the emission spectra of single probe molecules, and so unveil rich, nanoscale compositional heterogeneities in the membranes of live mammalian cells. With single-molecule displacement/diffusivity mapping (SMdM), we map out intracellular diffusivity with unprecedented spatial resolution and fidelity, and thus discover that diffusion in the mammalian cytoplasm and nucleus are both spatially heterogeneous at the nanoscale, and identify the net charge of the diffuser as a previously overlooked, key determinant of diffusion rate. In our recent efforts, we further turn spectral microscopy on its head by achieving exceptional multiplexing capabilities through demixing the fluorophore excitation spectrum. By adding remarkably rich functional dimensions to the already powerful super-resolution microscopy, we open up new ways to reveal fascinating local heterogeneities in living and chemical systems.