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Super-Resolution Live Cell Imaging of Subcellular Structures

Super-Resolution Live Cell Imaging of Subcellular Structures

Published on: January 13, 2021

Fluorescent proteins for live-cell imaging with super-resolution.

Karin Nienhaus¹, G Ulrich Nienhaus

¹Institute of Applied Physics and Center for Functional Nanostructures (CFN), Karlsruhe Institute of Technology (KIT), Wolfgang-Gaede-Straβe 1, 76131 Karlsruhe, Germany.

Chemical Society Reviews

|September 24, 2013

Summary

Fluorescent proteins (FPs) are vital for live imaging. Recent advances focus on photoactivatable, photoconvertible, and photoswitchable FPs for super-resolution microscopy and patterned illumination techniques.

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Area of Science:

Biochemistry
Molecular Biology
Microscopy

Background:

Fluorescent proteins (FPs) from the GFP family are essential genetic markers for live cell, tissue, and organism imaging.
Natural FPs have been extensively engineered to enhance their properties as genetically encoded markers.

Purpose of the Study:

To review recent developments in fluorescent protein engineering.
To highlight photoactivatable, photoconvertible, and photoswitchable FPs for advanced imaging techniques.

Main Methods:

Review of literature on FP isolation and engineering.
Focus on light-controlled FP activation mechanisms (photoactivation, photoconversion, photoswitching).
Discussion of FP applications in super-resolution microscopy (PALM, RESOLFT) and pulse-chase experiments.

Main Results:

Photoactivatable FPs enable regional marking and super-resolution imaging.
Photoconvertible FPs are suitable for localization-based super-resolution microscopy (PALM).
Photoswitchable FPs allow reversible activation for patterned illumination microscopy (RESOLFT), with recent variants showing improved switching fatigue resistance.

Conclusions:

Significant progress has been made in FP engineering for live imaging over two decades.
Further improvements in FP properties are still needed for optimal live-cell imaging applications.
Specialized FPs are crucial for emerging super-resolution and patterned illumination microscopy techniques.