Related Experiment Video
Updated: Jan 2, 2026
12:51
Simultaneous Multicolor Imaging of Biological Structures with Fluorescence Photoactivation Localization Microscopy
Published on: December 9, 2013
9.3K
Localization microscopy at doubled precision with patterned illumination
Jelmer Cnossen1,2, Taylor Hinsdale1, Rasmus Ø Thorsen1
1Department of Imaging Physics, Delft University of Technology, Delft, the Netherlands.
Nature Methods
|December 11, 2019
Summary
We enhanced single-molecule localization precision using widefield imaging, achieving a two-fold improvement over standard methods. This breakthrough improves super-resolution microscopy for biological imaging applications.
Area of Science:
- Super-resolution microscopy
- Biophysics
- Cell biology
Background:
- MINFLUX microscopy provides high precision but has a limited field of view.
- Extracting precise localization information from widefield images remains a challenge.
Purpose of the Study:
- To develop a method for enhanced single-molecule localization precision within a micrometer-sized field of view.
- To improve super-resolution imaging capabilities for biological samples.
Main Methods:
- Combined centroid estimation with illumination pattern-induced photon count variations.
- Utilized a conventional widefield imaging setup.
- Applied DNA-PAINT and STORM imaging techniques on DNA-origami nanostructures and cellular tubulin.
Main Results:
- Achieved a near two-fold improvement in localization precision compared to standard methods.
- Demonstrated enhanced precision with equivalent photon counts.
- Successfully applied the method to both purified nanostructures and complex cellular environments.
Conclusions:
- The developed method significantly enhances single-molecule localization precision in widefield microscopy.
- This approach overcomes field-of-view limitations of techniques like MINFLUX.
- Offers a promising advancement for high-resolution biological imaging.
More Related Videos
Related Concept Videos
01:37Super-resolution Fluorescence Microscopy
12.1K
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...
12.1K
01:29Two-Dimensional Microscopy in Microbiology
978
Two-dimensional (2D) microscopy encompasses a range of optical techniques that capture images within a single focal plane, offering detailed representations of microscopic structures. These techniques are essential in biological and medical research, enabling the visualization of cellular and subcellular structures with different levels of contrast and specificity.There are several major types of 2D microscopy, each with strengths and applications.Bright-Field MicroscopyBright-field microscopy...
978
01:16Confocal Fluorescence Microscopy
19.8K
Confocal microscopy is an advanced microscopic technique. The prime advantage of the confocal microscope over other microscopy techniques is its ability to block the out-of-focus light from the illuminated samples using pinholes. It is widely used with fluorescence optics to obtain high-resolution, sharp contrast images. Unlike optical microscopes, confocal microscopes use a focused beam of light laser to scan the entire sample surface at different z-planes. These microscopes are, therefore,...
19.8K
01:28Three-Dimensional Microscopy in Microbiology
715
Three-dimensional imaging techniques are essential in cell biology, allowing researchers to visualize intricate cellular structures with high resolution. Two prominent methods, Differential Interference Contrast Microscopy (DIC) and Confocal Scanning Laser Microscopy (CSLM), provide distinct advantages for imaging live and thick specimens, respectively.Differential Interference Contrast MicroscopyDIC microscopy enhances contrast in transparent, unstained samples by converting phase...
715