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Updated: Jun 16, 2025

A Guide to Structured Illumination TIRF Microscopy at High Speed with Multiple Colors

A Guide to Structured Illumination TIRF Microscopy at High Speed with Multiple Colors

Published on: May 30, 2016

Two-Dimensional Nonlinear Structured Illumination Microscopy with rsEGFP2.

Shaoheng Li¹, Ryo Tamura², Kota Banzai²

¹School of Electrical and Computer Engineering, University of Georgia, Athens, GA, USA.

Biorxiv : the Preprint Server for Biology

|June 4, 2025

Summary

This summary is machine-generated.

Nonlinear SIM (NSIM) microscopy achieves sub-80 nm resolution for live-cell actin imaging. This technique uses switchable fluorescent proteins and patterned depletion illumination for enhanced nanoscale detail.

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Implementation of a Nonlinear Microscope Based on Stimulated Raman Scattering

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Last Updated: Jun 16, 2025

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Implementation of a Nonlinear Microscope Based on Stimulated Raman Scattering

Implementation of a Nonlinear Microscope Based on Stimulated Raman Scattering

Published on: July 6, 2019

Area of Science:

Biophysics
Cell Biology
Microscopy

Background:

Superresolution microscopy offers nanoscale imaging of cellular structures and dynamics.
Structured illumination microscopy (SIM) is suitable for live-cell imaging.
Linear SIM provides a resolution of approximately 100 nm.

Purpose of the Study:

To achieve higher resolution live-cell imaging beyond linear SIM limitations.
To demonstrate the efficacy of nonlinear SIM (NSIM) for subcellular imaging.
To visualize dynamic cellular processes at the nanoscale.

Main Methods:

Utilized nonlinear SIM (NSIM) with reversibly switchable fluorescent proteins (rsEGFP2).
Employed patterned depletion illumination (PD) to generate nonlinear fluorescent response.
Performed 2D imaging of actin cytoskeleton in live U2OS cells.

Main Results:

Achieved sub-80 nm resolution in 2D imaging.
Successfully visualized nanoscale details of actin structures in live cells.
Demonstrated the capability of PD-NSIM for high-resolution live-cell dynamics.

Conclusions:

PD-NSIM significantly enhances resolution in live-cell superresolution microscopy.
This method allows for detailed observation of subcellular structures and dynamics.
NSIM offers a promising approach for advancing nanoscale biological imaging.