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Cryogenic Far-Field Fluorescence Nanoscopy: Evaluation with DNA Origami
Taku Furubayashi1, Keita Ishida1, Eiji Nakata2
1Department of Physics, Tokyo Institute of Technology, Meguro, Tokyo 152-8550, Japan.
The Journal of Physical Chemistry. B
|August 14, 2020
Summary
Researchers achieved 1-nanometer spatial accuracy in localizing individual fluorophores using DNA origami scaffolds at cryogenic temperatures. This advancement significantly improves precision in super-resolution microscopy by minimizing photobleaching and immobilizing fluorophores.
Area of Science:
- Nanotechnology
- Super-resolution microscopy
- Biophysics
Background:
- Achieving high spatial accuracy in super-resolution microscopy is crucial for understanding molecular mechanisms.
- Cryogenic temperatures offer benefits like fluorophore immobilization and reduced photobleaching, but can introduce systematic errors.
Purpose of the Study:
- To demonstrate far-field fluorescence localization nanoscopy with 1-nanometer accuracy at 1.8 K.
- To investigate and correct for systematic errors in cryogenic super-resolution imaging.
Main Methods:
- Utilized DNA origami as a 1-nanometer-accurate scaffold for immobilizing red and near-infrared fluorophores.
- Performed far-field fluorescence microscopy at cryogenic temperatures (1.8 K).
- Developed a 3D localization method to correct for dipole-orientation effects.
Main Results:
- Achieved lateral spatial precision of 1 nm under cryogenic conditions.
- Identified a lateral shift in fluorophore centroids due to dipole-orientation effects.
- Corrected for systematic errors, achieving a lateral spatial accuracy of 4 nm (standard error).
Conclusions:
- DNA origami scaffolds enable highly precise fluorophore localization at cryogenic temperatures.
- Correction for dipole-orientation effects is essential for accurate 3D super-resolution imaging.
- This technique advances the capability to visualize nanoscale structures with unprecedented accuracy.
