scholarly journals Dynamic Structure of Yeast Septin by Fast Fluctuation-Enhanced Structured Illumination Microscopy

2021 ◽  
Vol 9 (11) ◽  
pp. 2255
Author(s):  
Longfang Yao ◽  
Li Zhang ◽  
Liwen Chen ◽  
Xingyu Gong ◽  
Jiahui Zhong ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Super Resolution ◽  
Dynamic Structure ◽  
Frame Rate ◽  
Yeast Cells ◽  
Order Structure ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Division Site ◽  
Division Process

When Saccharomyces cerevisiae divides, a structure composed of different septin proteins arranged according to a certain rule is formed at the cell division site. The structure undergoes multiple remodeling stages during the cell cycle, thus guiding the yeast cells to complete the entire division process. Although the higher-order structure of septins can be determined using electron microscopy, the septin’s dynamic processes are poorly understood because of limitations in living cell super-resolution imaging technology. Herein, we describe a high lateral resolution and temporal resolution technique, known as fast fluctuation-enhanced structured illumination microscopy (fFE-SIM), which more than doubles the SIM resolution at a frame rate of 38 Hz in living cells. This allows a highly dynamic and sparse septin structure to be observed in Saccharomyces cerevisiae.

scholarly journals Extending resolution of structured illumination microscopy with sparse deconvolution

2021 ◽  
Author(s):  
Weisong Zhao ◽  
Shiqun Zhao ◽  
Liuju Li ◽  
Xiaoshuai Huang ◽  
Shijia Xing ◽  
...  
Keyword(s):  
Signal To Noise Ratio ◽  
Three Dimensional ◽  
Super Resolution ◽  
Live Cell ◽  
Frame Rate ◽  
Live Cells ◽  
Photon Flux ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Spatiotemporal Resolution

Abstract The spatial resolutions of live-cell super-resolution microscopes are limited by the maximum collected photon flux. Taking advantage of a priori knowledge of the sparsity and continuity of biological structures, we develop a deconvolution algorithm that further extends the resolution of super-resolution microscopes under the same photon budgets by nearly twofold. As a result, sparse structured illumination microscopy (Sparse-SIM) achieves ~60 nm resolution at a 564 Hz frame rate, allowing it to resolve intricate structural intermediates, including small vesicular fusion pores, ring-shaped nuclear pores formed by different nucleoporins, and relative movements between the inner and outer membranes of mitochondria in live cells. Likewise, sparse deconvolution can be used to increase the three-dimensional resolution and contrast of spinning-disc confocal-based SIM (SD-SIM), and operates under conditions with the insufficient signal-to-noise-ratio, all of which allows routine four-color, three-dimensional, ~90 nm resolution live-cell super-resolution imaging. Overall, sparse deconvolution may be a general tool to push the spatiotemporal resolution limits of live-cell fluorescence microscopy.

scholarly journals Applications of Super Resolution Expansion Microscopy in Yeast

2021 ◽  
Vol 9 ◽  
Author(s):  
Liwen Chen ◽  
Longfang Yao ◽  
Li Zhang ◽  
Yiyan Fei ◽  
Lan Mi ◽  
...  
Keyword(s):  
Super Resolution ◽  
Imaging Study ◽  
Yeast Cells ◽  
Nuclear Pore ◽  
Preparation Technique ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Resolution Imaging ◽  
Conventional Microscopy ◽  
Related Proteins

Super-resolution microscopy includes multiple techniques in optical microscopy that enable sub-diffraction resolution fluorescence imaging of cellular structures. Expansion microscopy (EXM) is a method of physical expansion to obtain super-resolution images of a biological sample on conventional microscopy. We present images of yeast organelles, applying the combination of super-resolution and ExM techniques. When preparing pre-expanded samples, conventional methods lead to breakage of dividing yeast cells and difficulties in studying division-related proteins. Here, we describe an improved sample preparation technique that avoids such damage. ExM in combination with Airyscan and structured illumination microscopy (SIM) collected sub-cellular structural images of nuclear pore complex, septin, and a-tubulin in yeast. Our method of expansion in yeast is well-suited for super-resolution imaging study of yeast.

Live, Video-Rate Super-Resolution Microscopy Using Structured Illumination and Rapid GPU-Based Parallel Processing

2011 ◽  
Vol 17 (2) ◽  
pp. 191-196 ◽  
Author(s):  
Jonathan Lefman ◽  
Keana Scott ◽  
Stephan Stranick
Keyword(s):  
Parallel Processing ◽  
Real Time ◽  
Super Resolution ◽  
Fold Increase ◽  
Frame Rate ◽  
Processing Unit ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Multiprocessor Computer ◽  
Computational Overhead

AbstractStructured illumination fluorescence microscopy is a powerful super-resolution method that is capable of achieving a resolution below 100 nm. Each super-resolution image is computationally constructed from a set of differentially illuminated images. However, real-time application of structured illumination microscopy (SIM) has generally been limited due to the computational overhead needed to generate super-resolution images. Here, we have developed a real-time SIM system that incorporates graphic processing unit (GPU) based in-line parallel processing of raw/differentially illuminated images. By using GPU processing, the system has achieved a 90-fold increase in processing speed compared to performing equivalent operations on a multiprocessor computer—the total throughput of the system is limited by data acquisition speed, but not by image processing. Overall, more than 350 raw images (16-bit depth, 512 × 512 pixels) can be processed per second, resulting in a maximum frame rate of 39 super-resolution images per second. This ultrafast processing capability is used to provide immediate feedback of super-resolution images for real-time display. These developments are increasing the potential for sophisticated super-resolution imaging applications.

scholarly journals Two-dimensional TIRF-SIM–traction force microscopy (2D TIRF-SIM-TFM)

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Liliana Barbieri ◽  
Huw Colin-York ◽  
Kseniya Korobchevskaya ◽  
Di Li ◽  
Deanna L. Wolfson ◽  
...  
Keyword(s):  
Resolution Enhancement ◽  
Traction Force ◽  
Super Resolution ◽  
Traction Force Microscopy ◽  
Two Dimensional ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Force Microscopy ◽  
Health And Disease ◽  
Spatio Temporal

AbstractQuantifying small, rapidly evolving forces generated by cells is a major challenge for the understanding of biomechanics and mechanobiology in health and disease. Traction force microscopy remains one of the most broadly applied force probing technologies but typically restricts itself to slow events over seconds and micron-scale displacements. Here, we improve >2-fold spatially and >10-fold temporally the resolution of planar cellular force probing compared to its related conventional modalities by combining fast two-dimensional total internal reflection fluorescence super-resolution structured illumination microscopy and traction force microscopy. This live-cell 2D TIRF-SIM-TFM methodology offers a combination of spatio-temporal resolution enhancement relevant to forces on the nano- and sub-second scales, opening up new aspects of mechanobiology to analysis.

scholarly journals Double moiré localized plasmon structured illumination microscopy

Nanophotonics ◽  
2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Ruslan Röhrich ◽  
A. Femius Koenderink
Keyword(s):  
Near Field ◽  
Super Resolution ◽  
Reconstruction Algorithm ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Experimental Realization ◽  
Spatial Frequencies ◽  
Wide Range ◽  
Localized Plasmon ◽  
Plasmonic Grating

AbstractStructured illumination microscopy (SIM) is a well-established fluorescence imaging technique, which can increase spatial resolution by up to a factor of two. This article reports on a new way to extend the capabilities of structured illumination microscopy, by combining ideas from the fields of illumination engineering and nanophotonics. In this technique, plasmonic arrays of hexagonal symmetry are illuminated by two obliquely incident beams originating from a single laser. The resulting interference between the light grating and plasmonic grating creates a wide range of spatial frequencies above the microscope passband, while still preserving the spatial frequencies of regular SIM. To systematically investigate this technique and to contrast it with regular SIM and localized plasmon SIM, we implement a rigorous simulation procedure, which simulates the near-field illumination of the plasmonic grating and uses it in the subsequent forward imaging model. The inverse problem, of obtaining a super-resolution (SR) image from multiple low-resolution images, is solved using a numerical reconstruction algorithm while the obtained resolution is quantitatively assessed. The results point at the possibility of resolution enhancements beyond regular SIM, which rapidly vanishes with the height above the grating. In an initial experimental realization, the existence of the expected spatial frequencies is shown and the performance of compatible reconstruction approaches is compared. Finally, we discuss the obstacles of experimental implementations that would need to be overcome for artifact-free SR imaging.

scholarly journals High-fidelity structured illumination microscopy by point-spread-function engineering

2021 ◽  
Vol 10 (1) ◽  
Author(s):  
Gang Wen ◽  
Simin Li ◽  
Linbo Wang ◽  
Xiaohu Chen ◽  
Zhenglong Sun ◽  
...  
Keyword(s):  
Point Spread Function ◽  
Super Resolution ◽  
Reconstruction Algorithm ◽  
High Fidelity ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Imaging Tool ◽  
Point Spread ◽  
Spread Function ◽  
Point Spread Function Engineering

AbstractStructured illumination microscopy (SIM) has become a widely used tool for insight into biomedical challenges due to its rapid, long-term, and super-resolution (SR) imaging. However, artifacts that often appear in SIM images have long brought into question its fidelity, and might cause misinterpretation of biological structures. We present HiFi-SIM, a high-fidelity SIM reconstruction algorithm, by engineering the effective point spread function (PSF) into an ideal form. HiFi-SIM can effectively reduce commonly seen artifacts without loss of fine structures and improve the axial sectioning for samples with strong background. In particular, HiFi-SIM is not sensitive to the commonly used PSF and reconstruction parameters; hence, it lowers the requirements for dedicated PSF calibration and complicated parameter adjustment, thus promoting SIM as a daily imaging tool.

The FDTD Analysis of Near-Field Response for Microgroove Structure With Standing Wave Illumination for the Realization of Coherent Structured Illumination Microscopy

2021 ◽  
Author(s):  
Yizhao Guan ◽  
Hiromasa Kume ◽  
Shotaro Kadoya ◽  
Masaki Michihata ◽  
Satoru Takahashi
Keyword(s):  
Standing Wave ◽  
Incident Angle ◽  
Near Field ◽  
Super Resolution ◽  
Structured Illumination ◽  
Field Phase ◽  
Structured Illumination Microscopy ◽  
Depth Measurement ◽  
Field Response ◽  
Depth Dependency

Abstract Microstructures are widely used in the manufacture of functional surfaces. An optical-based super-resolution, non-invasive method is preferred for the inspection of surfaces with massive microstructures. The Structured Illumination Microscopy (SIM) uses standing-wave illumination to reach optical super-resolution. Recently, coherent SIM is being studied. It can obtain not only the super-resolved intensity distribution but also the phase and amplitude distribution of the sample surface beyond the diffraction limit. By analysis of the phase-depth dependency, the depth measurement for microgroove structures with coherent SIM is expected. FDTD analysis is applied for observing the near-field response of microgroove under the standing-wave illumination. The near-field phase shows depth dependency in this analysis. Moreover, the effects from microgroove width, the incident angle, and the relative position between the standing-wave peak and center of the microgroove are investigated. It is found the near-field phase change can measure depth until 200 nm (aspect ratio 1) with an error of up to 20.4 nm in the case that the microgroove width is smaller than half of the wavelength.

Super-Resolution Imaging in Fluid Mechanics Using New Illumination Approaches

2020 ◽  
Vol 52 (1) ◽  
pp. 369-393
Author(s):  
Minami Yoda
Keyword(s):  
Fluid Mechanics ◽  
Total Internal Reflection ◽  
Imaging Techniques ◽  
Super Resolution ◽  
Internal Reflection ◽  
Interfacial Flows ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Entire Volume ◽  
Resolution Imaging

Quantifying submillimeter flows using optical diagnostic techniques is often limited by a lack of spatial resolution and optical access. This review discusses two super-resolution imaging techniques, structured illumination microscopy and total internal reflection fluorescence or microscopy, which can visualize bulk and interfacial flows, respectively, at spatial resolutions below the classic diffraction limits. First, we discuss the theory and applications of structured illumination for optical sectioning, i.e., imaging a thin slice of a flow illuminated over its entire volume. Structured illumination can be used to visualize the interior of multiphase flows such as sprays by greatly reducing secondary scattering. Second, the theory underlying evanescent waves is introduced, followed by a review of how total internal reflection microscopy has been used to visualize interfacial flows over the last 15 years. Both techniques, which are starting to be used in fluid mechanics, could significantly improve quantitative imaging of microscale and macroscale flows.

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