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Related Experiment Video

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Conventional BODIPY Conjugates for Live-Cell Super-Resolution Microscopy and Single-Molecule Tracking

Conventional BODIPY Conjugates for Live-Cell Super-Resolution Microscopy and Single-Molecule Tracking

Published on: June 8, 2020

3D single-molecule super-resolution microscopy with a tilted light sheet.

Anna-Karin Gustavsson^1,2, Petar N Petrov¹, Maurice Y Lee^1,3

¹Department of Chemistry, Stanford University, Stanford, CA, 94305, USA.

Nature Communications

|January 11, 2018

Summary

This summary is machine-generated.

Tilted light sheet microscopy with 3D point spread functions (TILT3D) enables precise 3D super-resolution imaging in thick cells. This method achieves tens of nanometer localization precision for biological structures.

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Area of Science:

Microscopy
Biophysics
Optical Imaging

Background:

3D super-resolution microscopy is crucial for visualizing cellular structures.
Existing methods often struggle with thick samples and achieving high axial precision.

Purpose of the Study:

To develop a novel 3D super-resolution microscopy technique for thick biological samples.
To achieve high-precision single-molecule localization and imaging in mammalian cells.

Main Methods:

Implementation of tilted light sheet illumination with long axial range point spread functions (PSFs).
Utilized double-helix PSF for single-molecule detection and tetrapod PSFs for fiducial bead tracking.
Developed a system (TILT3D) built on a standard inverted microscope with minimal custom parts.

Main Results:

Demonstrated low-background, 3D super-localization of single molecules in thick cells.
Achieved tens of nanometers localization precision throughout mammalian cells.
Successfully imaged mitochondria and the nuclear lamina with high resolution.

Conclusions:

TILT3D offers a simple and flexible approach to 3D super-resolution imaging.
The method excels in thick samples, overcoming limitations of thin light sheets.
Validated for high-precision imaging of cellular ultrastructures in mammalian cells.