Biology subjects

Pott, E. D.

Publications and source records attributed to Pott, E. D..

3 recordsLinked to original sources

bioRxiv2025

Fast Hyperspectral and Super-Resolved Mapping of Lipid Membrane Polarity with Single-Molecule Sensitivity

Cell membranes display nanoscale heterogeneity in lipid composition and organization that regulates vital biological processes yet remains challenging to resolve with conventional imaging. We introduce spectral phasor single molecule localization microscopy (SP-SMLM), a hyperspectral and super-resolution method that combines wavefront-like optical filtering with single molecule imaging for simultaneous spatial and spectral analysis. A lab-built three-channel imager with sine/cosine filters encodes emission spectra of single molecules into the phasor space, enabling high-throughput, high-SNR mapping of membrane polarity at sub-50 nm spatial and 15-sec temporal resolutions. Through simulation, we validate that the phasor angle correlates with the spectral mean for single dye molecules. When applied to Nile Red-stained COS-7 cells, SP-SMLM revealed organelle-specific polarity differences and dynamic remodeling of lipid composition within live cells. The methods hyperspectral capability, rapid acquisition, and compatibility with 2D/3D imaging platforms position SP-SMLM as a powerful tool for studying membrane heterogeneity and dynamics in live cells.

biophysics↗

bioRxiv2025

High Throughput Hyperspectral and Multiplexed Super-Resolution Fluorescence Imaging by SP-STORM

Simultaneous determination of spatial location and spectral color of single molecules at large molecular density with high throughput was achieved by combining single molecule photoswitching and in-hardware transformation based spectral phasor analysis. The method, named spectral phasor enabled stochastic optical reconstruction microscopy (SP-STORM), achieved simultaneous super-resolution imaging of five subcellular structures with minimum crosstalk for the first time. The high throughput feature of SP-STORM enables these subcellular structures to be readily resolved in about one minute, which is more than a magnitude faster than other multiplexing single molecule localization microscopy techniques. The concept of SP-STORM is also compatible with and can be readily applicable to other super-resolution microscopy.

biophysics↗

bioRxiv2024

Spectrally Resolved Localization Microscopy with Ultrahigh-Throughput

Single-molecule localization microscopy (SMLM) has become a strong technique in the toolbox of chemists, biologists, physicists, and engineers in recent years for its unique ability to resolve characteristic features quickly and accurately in complex environments at the nanoscopic level. Multicolor super-resolution imaging has seen the greatest advancement among SMLM techniques, drastically improving the differentiation ability of nanostructures beyond the diffraction limit and increasing the resolution with which previously unresolvable structures are studied. However, current multicolor SMLM methodologies present low spatial resolution and throughput and require complex optical systems. Here, we overcome these drawbacks by developing an ultrahigh-throughput SMLM methodology that allows for ultrahigh throughput multicolor imaging at the nanoscopic level using a color glass filter. Our methodology can readily distinguish fluorophores of close spectral emission and achieves sub-10 nm localization and sub-5nm spectral precisions.

biophysics↗