scholarly journals Quantification of cristae architecture reveals time-dependent characteristics of individual mitochondria

2020 ◽  
Vol 3 (7) ◽  
pp. e201900620
Author(s):  
Mayuko Segawa ◽  
Dane M Wolf ◽  
Nan W Hultgren ◽  
David S Williams ◽  
Alexander M van der Bliek ◽  
...  
Keyword(s):  
Machine Learning ◽  
Mitochondrial Fission ◽  
Live Cell ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Fusion Event ◽  
New Strategy ◽  
Function Relationship ◽  
Relationship Of ◽  
Standard Techniques

Recent breakthroughs in live-cell imaging have enabled visualization of cristae, making it feasible to investigate the structure–function relationship of cristae in real time. However, quantifying live-cell images of cristae in an unbiased way remains challenging. Here, we present a novel, semi-automated approach to quantify cristae, using the machine-learning Trainable Weka Segmentation tool. Compared with standard techniques, our approach not only avoids the bias associated with manual thresholding but more efficiently segments cristae from Airyscan and structured illumination microscopy images. Using a cardiolipin-deficient cell line, as well as FCCP, we show that our approach is sufficiently sensitive to detect perturbations in cristae density, size, and shape. This approach, moreover, reveals that cristae are not uniformly distributed within the mitochondrion, and sites of mitochondrial fission are localized to areas of decreased cristae density. After a fusion event, individual cristae from the two mitochondria, at the site of fusion, merge into one object with distinct architectural values. Overall, our study shows that machine learning represents a compelling new strategy for quantifying cristae in living cells.

Cost-Effective Live Cell Structured Illumination Microscopy with Video-Rate Imaging

ACS Photonics ◽  
2021 ◽  
Author(s):  
Alice Sandmeyer ◽  
Mario Lachetta ◽  
Hauke Sandmeyer ◽  
Wolfgang Hübner ◽  
Thomas Huser ◽  
...  
Keyword(s):  
Cost Effective ◽  
Live Cell ◽  
Structured Illumination ◽  
Structured Illumination Microscopy

scholarly journals Rapid bacteria identification using structured illumination microscopy and machine learning

2017 ◽  
Vol 11 (01) ◽  
pp. 1850007 ◽  
Author(s):  
Yingchuan He ◽  
Weize Xu ◽  
Yao Zhi ◽  
Rohit Tyagi ◽  
Zhe Hu ◽  
...  
Keyword(s):  
Machine Learning ◽  
Optical Microscopy ◽  
Pathogenic Bacteria ◽  
Morphological Features ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Standard Pattern ◽  
Bacteria Identification ◽  
Detection And Identification ◽  
Combined Use

Traditionally, optical microscopy is used to visualize the morphological features of pathogenic bacteria, of which the features are further used for the detection and identification of the bacteria. However, due to the resolution limitation of conventional optical microscopy as well as the lack of standard pattern library for bacteria identification, the effectiveness of this optical microscopy-based method is limited. Here, we reported a pilot study on a combined use of Structured Illumination Microscopy (SIM) with machine learning for rapid bacteria identification. After applying machine learning to the SIM image datasets from three model bacteria (including Escherichia coli, Mycobacterium smegmatis, and Pseudomonas aeruginosa), we obtained a classification accuracy of up to 98%. This study points out a promising possibility for rapid bacterial identification by morphological features.

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 Advances in High-Speed Structured Illumination Microscopy

2021 ◽  
Vol 9 ◽  
Author(s):  
Tianyu Zhao ◽  
Zhaojun Wang ◽  
Tongsheng Chen ◽  
Ming Lei ◽  
Baoli Yao ◽  
...  
Keyword(s):  
Live Cell Imaging ◽  
High Speed ◽  
Cell Imaging ◽  
Super Resolution ◽  
Live Cell ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Light Power ◽  
Recent Developments ◽  
Super Resolution Microscopy

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.

scholarly journals Mitochondrial RNA granules are fluid condensates, positioned by membrane dynamics

2019 ◽  
Author(s):  
Timo Rey ◽  
Sofia Zaganelli ◽  
Emilie Cuillery ◽  
Jean-Claude Martinou ◽  
Suliana Manley
Keyword(s):  
Dynamic Properties ◽  
Mitochondrial Fission ◽  
Super Resolution ◽  
Membrane Dynamics ◽  
Cellular Respiration ◽  
Mitochondrial Rna ◽  
Structured Illumination ◽  
Inner Mitochondrial Membrane ◽  
Structured Illumination Microscopy ◽  
Rna Granules

Mitochondria contain the genetic information and expression machinery to produce proteins essential for cellular respiration. Within the mitochondrial matrix, newly synthesized RNA, RNA processing proteins, and mitoribosome assembly factors are known to form punctate subcompartments referred to as mitochondrial RNA granules (MRGs) 1–3. Despite their proposed role in regulating gene expression, little is known about the structural and dynamic properties of MRGs. We investigated the organization of MRGs using fluorescence super-resolution localization microscopy and correlative electron microscopy techniques, obtaining ultrastructural details of their internal architecture. We find that MRGs are organized into nanoscale RNA cores surrounded by a protein shell. Using live-cell super-resolution structured illumination microscopy and photobleaching perturbations, we reveal that MRGs undergo fusion and rapidly exchange components, consistent with liquid-liquid phase separation (LLPS). Furthermore, MRGs associate with the inner mitochondrial membrane and their fusion coincides with membrane remodeling. Inhibition of mitochondrial fission leads to an aberrant distribution of MRGs into concentrated pockets, where they remain as distinct individual units despite their close apposition. Together, our results reveal a role for LLPS in concentrating RNA and its processing proteins into MRGs, which are positioned along mitochondria by membrane dynamics.

scholarly journals Mitochondrial membrane tension governs fission

2018 ◽  
Author(s):  
Dora Mahecic ◽  
Lina Carlini ◽  
Tatjana Kleele ◽  
Adai Colom ◽  
Antoine Goujon ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Mitochondrial Membrane ◽  
Mitochondrial Fission ◽  
Outer Mitochondrial Membrane ◽  
Membrane Tension ◽  
Structured Illumination ◽  
Bending Energy ◽  
Structured Illumination Microscopy ◽  
Cellular Factors ◽  
Tension Sensor

AbstractDuring mitochondrial fission, key molecular and cellular factors assemble on the outer mitochondrial membrane, where they coordinate to generate constriction. Constriction sites can eventually divide, or reverse upon disassembly of the machinery. However, a role for membrane tension in mitochondrial fission, although speculated, has remained undefined. We captured the dynamics of constricting mitochondria in mammalian cells using live-cell structured illumination microscopy (SIM). By analyzing the diameters of tubules that emerge from mitochondria and implementing a fluorescence lifetime-based mitochondrial membrane tension sensor, we discovered that mitochondria are indeed under tension. Under perturbations that reduce mitochondrial tension, constrictions initiate at the same rate, but are less likely to divide. We propose a model based on our estimates of mitochondrial membrane tension and bending energy in living cells which accounts for the observed probability distribution for mitochondrial constrictions to divide.

Evaluating resolution in live cell structured illumination microscopy

2019 ◽  
Author(s):  
Jakub Pospisil ◽  
Karel Fliegel ◽  
Jan Švihlík ◽  
Miloš Klíma
Keyword(s):  
Live Cell ◽  
Structured Illumination ◽  
Structured Illumination Microscopy

scholarly journals Motion artefact detection in structured illumination microscopy for live cell imaging

2016 ◽  
Vol 24 (19) ◽  
pp. 22121 ◽  
Author(s):  
Ronny Förster ◽  
Kai Wicker ◽  
Walter Müller ◽  
Aurélie Jost ◽  
Rainer Heintzmann
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Motion Artefact ◽  
Artefact Detection

Live Cell Imaging With Spatial Light Modulator-based Optical Sectioning Structured Illumination Microscopy

2014 ◽  
Vol 20 (S3) ◽  
pp. 388-389
Author(s):  
Zdeněk Švindrych ◽  
Pavel Křížek ◽  
Evgeny Smirnov ◽  
Martin Ovesný ◽  
Josef Borkovec ◽  
...  
Keyword(s):  
Live Cell Imaging ◽  
Spatial Light Modulator ◽  
Cell Imaging ◽  
Live Cell ◽  
Structured Illumination ◽  
Optical Sectioning ◽  
Structured Illumination Microscopy ◽  
Light Modulator
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