scholarly journals Ultrafast superresolution fluorescence imaging with spinning disk confocal microscope optics

2015 ◽  
Vol 26 (9) ◽  
pp. 1743-1751 ◽  
Author(s):  
Shinichi Hayashi ◽  
Yasushi Okada

Most current superresolution (SR) microscope techniques surpass the diffraction limit at the expense of temporal resolution, compromising their applications to live-cell imaging. Here we describe a new SR fluorescence microscope based on confocal microscope optics, which we name the spinning disk superresolution microscope (SDSRM). Theoretically, the SDSRM is equivalent to a structured illumination microscope (SIM) and achieves a spatial resolution of 120 nm, double that of the diffraction limit of wide-field fluorescence microscopy. However, the SDSRM is 10 times faster than a conventional SIM because SR signals are recovered by optical demodulation through the stripe pattern of the disk. Therefore a single SR image requires only a single averaged image through the rotating disk. On the basis of this theory, we modified a commercial spinning disk confocal microscope. The improved resolution around 120 nm was confirmed with biological samples. The rapid dynamics of micro­tubules, mitochondria, lysosomes, and endosomes were observed with temporal resolutions of 30–100 frames/s. Because our method requires only small optical modifications, it will enable an easy upgrade from an existing spinning disk confocal to a SR microscope for live-cell imaging.

Photonics ◽  
2021 ◽  
Vol 8 (7) ◽  
pp. 275
Author(s):  
Herbert Schneckenburger ◽  
Verena Richter

A short overview on 3D live cell imaging is given. Relevant samples are described and various problems and challenges—including 3D imaging by optical sectioning, light scattering and phototoxicity—are addressed. Furthermore, enhanced methods of wide-field or laser scanning microscopy together with some relevant examples and applications are summarized. In the future one may profit from a continuous increase in microscopic resolution, but also from molecular sensing techniques in the nanometer range using e.g., non-radiative energy transfer (FRET).


2021 ◽  
Vol 120 (3) ◽  
pp. 179a
Author(s):  
Peter T. Brown ◽  
Rory Kruithoff ◽  
Gregory J. Seedorf ◽  
Douglas P. Shepherd

2020 ◽  
Vol 13 (4) ◽  
pp. 045002
Author(s):  
Tomu Suzuki ◽  
Shinji Kajimoto ◽  
Narufumi Kitamura ◽  
Mayumi Takano-Kasuya ◽  
Naoko Furusawa ◽  
...  

2021 ◽  
Vol 9 ◽  
Author(s):  
Tianyu Zhao ◽  
Zhaojun Wang ◽  
Tongsheng Chen ◽  
Ming Lei ◽  
Baoli Yao ◽  
...  

Super-resolution microscopy surpasses the diffraction limit to enable the observation of the fine details in sub-cellular structures and their dynamics in diverse biological processes within living cells. Structured illumination microscopy (SIM) uses a relatively low illumination light power compared with other super-resolution microscopies and has great potential to meet the demands of live-cell imaging. However, the imaging acquisition and reconstruction speeds limit its further applications. In this article, recent developments all targeted at improving the overall speed of SIM are reviewed. These comprise both hardware and software improvements, which include a reduction in the number of raw images, GPU acceleration, deep learning and the spatial domain reconstruction. We also discuss the application of these developments in live-cell imaging.


2011 ◽  
Vol 135 (2) ◽  
pp. 255-263
Author(s):  
Daniel B Schmolze ◽  
Clive Standley ◽  
Kevin E Fogarty ◽  
Andrew H Fischer

Abstract Context.—Advances in microscopy enable visualization of a broad range of new morphologic features. Objective.—To review and illustrate advances in microscopy with relevance to pathologists. Data Sources.—Literature review and new observations. Results.—Fluorescence microscopy enables multiantigen detection; allows novel optical-sectioning techniques, with some advantages compared to paraffin sectioning; and permits live-cell imaging. Live-cell imaging allows pathologists to move from a period when all diagnostic expertise was reliant on interpreting static images to a period when cellular dynamics can play a role in diagnosis. New techniques have bypassed by about 100-fold what had long been believed to be a limit to the resolution of light microscopy. Fluorescence resonance energy transfer (FRET) appears capable of visualizing diagnostically relevant molecular events in living or fixed cells that are immeasurable by other molecular techniques. We describe applications of 2-photon microscopy, FRET, structured illumination, and the subdiffraction techniques of near-field microscopy, photoactivated localization microscopy, stochastic optical reconstruction microscopy, and stimulated emission depletion microscopy. Conclusion.—New microscopy techniques present opportunities for pathologists to develop improved diagnostic tests.


2016 ◽  
Vol 24 (19) ◽  
pp. 22121 ◽  
Author(s):  
Ronny Förster ◽  
Kai Wicker ◽  
Walter Müller ◽  
Aurélie Jost ◽  
Rainer Heintzmann

2014 ◽  
Vol 20 (S3) ◽  
pp. 388-389
Author(s):  
Zdeněk Švindrych ◽  
Pavel Křížek ◽  
Evgeny Smirnov ◽  
Martin Ovesný ◽  
Josef Borkovec ◽  
...  

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