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Super-resolution Fluorescence Microscopy

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Super-resolution fluorescence microscopy (SRFM) provides a better resolution than conventional fluorescence microscopy by reducing the point spread function (PSF). PSF is the light intensity distribution from a point that causes it to appear blurred. Due to PSF, each fluorescing point appears bigger than its actual size, and it is the PSF interference of nearby fluorophores that causes the blurred image. Various approaches to achieving higher resolution through SRFM have recently been...

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Updated: Sep 24, 2025

Super-resolution Imaging of the Bacterial Division Machinery

Super-resolution Imaging of the Bacterial Division Machinery

Published on: January 21, 2013

Fluorogenic DNA-PAINT for faster, low-background super-resolution imaging.

Kenny K H Chung^1,2, Zhao Zhang^1,2, Phylicia Kidd¹

¹Department of Cell Biology, Yale University, New Haven, CT, USA.

|May 2, 2022

Summary

This summary is machine-generated.

Fluorogenic DNA-PAINT enhances super-resolution microscopy by reducing background noise and increasing imaging speed. This novel approach improves fluorescence and enables faster, 3D nanoscale imaging without optical sectioning.

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Last Updated: Sep 24, 2025

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Area of Science:

Microscopy
Biotechnology
Nanotechnology

Background:

DNA-based points accumulation for imaging in nanoscale topography (DNA-PAINT) is a super-resolution microscopy technique.
Existing DNA-PAINT methods are limited by high background noise and slow imaging speeds due to unbound fluorophores.

Purpose of the Study:

To develop an improved two-color fluorogenic DNA-PAINT method.
To address limitations of high background and slow imaging in DNA-PAINT.

Main Methods:

Designed self-quenching, single-stranded DNA imager probes conjugated with fluorophores and quenchers.
Engineered base pair mismatches between fluorogenic imager probes and docking strands.
Implemented two-color fluorogenic DNA-PAINT.

Main Results:

Achieved up to a 57-fold increase in fluorescence upon probe binding and unquenching.
Demonstrated a 26-fold increase in imaging speed compared to conventional DNA-PAINT.
Enabled three-dimensional super-resolution DNA-PAINT imaging without optical sectioning.

Conclusions:

Fluorogenic DNA-PAINT significantly enhances imaging speed and reduces background noise.
The new probe and docking strand designs improve fluorescence efficiency and binding kinetics.
This method facilitates advanced 3D super-resolution imaging at the nanoscale.