Recent advances in super-resolution fluorescence microscopy have led to ~10 nm spatial resolution and exciting new biology. We are developing new approaches to advance beyond the structural (shape) information offered by existing super-resolution methods, and reveal multidimensional information of intracellular <i>functional</i> parameters, including chemical polarity, diffusivity, and reactivity, with nanoscale resolution and single-molecule sensitivity. In particular, we show that the use of polarity-sensitive, solvatochromic fluorescence dyes with spectrally resolved super-resolution microscopy enables the probing of local chemical polarity at the nanometer scale. We thus reveal unexpectedly rich compositional and property heterogeneities for cell membranes and for surface adlayers. Meanwhile, using a conventional fluorescent protein, we recently developed a novel super-resolution method that enables the nanoscale mapping of intracellular diffusivity through determining the instantaneous displacements of single molecules diffusing freely inside the cell. We thus show that diffusion in the cytoplasm and in the nucleus to both be spatially heterogeneous at the nanoscale, and identify positive net charge as an overlooked key factor for intercellular diffusion slowdown. By adding remarkably rich <i>functional</i> dimensions to the already powerful super-resolution microscopy, we thus open up new ways to reveal fascinating local heterogeneities in live cells. Future work will benefit greatly from a synergy between continued advances in both instrumentation and fluorescence probe development.

Multifunctional super-resolution microscopy with solvatochromic and conventional fluorescent probes

2019 Fall National Meeting

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