Simultaneous spatiotemporal super-resolution and multi-parametric fluorescence microscopy

Jagadish Sankaran, Harikrushnan Balasubramanian, Wai Hoh Tang, Xue Wen Ng, Adrian Röllin and Thorsten Wohland ()
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Jagadish Sankaran: National University of Singapore
Harikrushnan Balasubramanian: National University of Singapore
Wai Hoh Tang: National University of Singapore
Xue Wen Ng: National University of Singapore
Adrian Röllin: National University of Singapore
Thorsten Wohland: National University of Singapore

Nature Communications, 2021, vol. 12, issue 1, 1-14

Abstract: Abstract Super-resolution microscopy and single molecule fluorescence spectroscopy require mutually exclusive experimental strategies optimizing either temporal or spatial resolution. To achieve both, we implement a GPU-supported, camera-based measurement strategy that highly resolves spatial structures (~100 nm), temporal dynamics (~2 ms), and molecular brightness from the exact same data set. Simultaneous super-resolution of spatial and temporal details leads to an improved precision in estimating the diffusion coefficient of the actin binding polypeptide Lifeact and corrects structural artefacts. Multi-parametric analysis of epidermal growth factor receptor (EGFR) and Lifeact suggests that the domain partitioning of EGFR is primarily determined by EGFR-membrane interactions, possibly sub-resolution clustering and inter-EGFR interactions but is largely independent of EGFR-actin interactions. These results demonstrate that pixel-wise cross-correlation of parameters obtained from different techniques on the same data set enables robust physicochemical parameter estimation and provides biological knowledge that cannot be obtained from sequential measurements.

Date: 2021
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DOI: 10.1038/s41467-021-22002-9

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