scholarly journals Low-Power Sonication Can Alter Extracellular Vesicle Size and Properties

Cells ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 2413
Author(s):  
Zubair Ahmed Nizamudeen ◽  
Rachael Xerri ◽  
Christopher Parmenter ◽  
Kiran Suain ◽  
Robert Markus ◽  
...  
Keyword(s):  
Low Power ◽  
Cultured Cells ◽  
Membrane Integrity ◽  
Extracellular Vesicle ◽  
Systematic Analysis ◽  
Vesicle Size ◽  
Stochastic Optical Reconstruction Microscopy ◽  
Transmission Electron ◽  
Power Setting ◽  
Optical Reconstruction

Low-power sonication is widely used to disaggregate extracellular vesicles (EVs) after isolation, however, the effects of sonication on EV samples beyond dispersion are unclear. The present study analysed the characteristics of EVs collected from mesenchymal stem cells (MSCs) after sonication, using a combination of transmission electron microscopy, direct stochastic optical reconstruction microscopy, and flow cytometry techniques. Results showed that beyond the intended disaggregation effect, sonication using the lowest power setting available was enough to alter the size distribution, membrane integrity, and uptake of EVs in cultured cells. These results point to the need for a more systematic analysis of sonication procedures to improve reproducibility in EV-based cellular experiments.

Dual responsive specifically labelled carbogenic fluorescent nanodots for super resolution and electron microscopy

Nanoscale ◽  
2019 ◽  
Vol 11 (14) ◽  
pp. 6561-6565 ◽  
Author(s):  
Navneet. C. Verma ◽  
Chethana Rao ◽  
Ashutosh Singh ◽  
Neha Garg ◽  
Chayan K. Nandi
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Single Molecule ◽  
Super Resolution ◽  
Dual Responsive ◽  
Stochastic Optical Reconstruction Microscopy ◽  
Transmission Electron ◽  
Optical Reconstruction

We introduce an orange emissive fluorescent nanodot for successful single molecule stochastic optical reconstruction microscopy (STORM), super resolution radial fluctuation (SRRF) microscopy and transmission electron microscopy (TEM).

scholarly journals Blinking statistics and molecular counting in direct stochastic reconstruction microscopy (dSTORM)

2021 ◽  
Author(s):  
Lekha Patel ◽  
David Williamson ◽  
Dylan M Owen ◽  
Edward A K Cohen
Keyword(s):  
Probability Distribution ◽  
Single Molecule ◽  
Immunological Synapse ◽  
Training Data ◽  
Supplementary Information ◽  
Cellular Structures ◽  
Exact Probability ◽  
Stochastic Optical Reconstruction Microscopy ◽  
Exact Probability Distribution ◽  
Optical Reconstruction

Abstract Motivation Many recent advancements in single-molecule localization microscopy exploit the stochastic photoswitching of fluorophores to reveal complex cellular structures beyond the classical diffraction limit. However, this same stochasticity makes counting the number of molecules to high precision extremely challenging, preventing key insight into the cellular structures and processes under observation. Results Modelling the photoswitching behaviour of a fluorophore as an unobserved continuous time Markov process transitioning between a single fluorescent and multiple dark states, and fully mitigating for missed blinks and false positives, we present a method for computing the exact probability distribution for the number of observed localizations from a single photoswitching fluorophore. This is then extended to provide the probability distribution for the number of localizations in a direct stochastic optical reconstruction microscopy experiment involving an arbitrary number of molecules. We demonstrate that when training data are available to estimate photoswitching rates, the unknown number of molecules can be accurately recovered from the posterior mode of the number of molecules given the number of localizations. Finally, we demonstrate the method on experimental data by quantifying the number of adapter protein linker for activation of T cells on the cell surface of the T-cell immunological synapse. Availability and implementation Software and data available at https://github.com/lp1611/mol_count_dstorm. Supplementary information Supplementary data are available at Bioinformatics online.

scholarly journals Molecular organization of mammalian meiotic chromosome axis revealed by expansion STORM microscopy

2019 ◽  
Vol 116 (37) ◽  
pp. 18423-18428 ◽  
Author(s):  
Huizhong Xu ◽  
Zhisong Tong ◽  
Qing Ye ◽  
Tengqian Sun ◽  
Zhenmin Hong ◽  
...  
Keyword(s):  
Meiotic Chromosome ◽  
Molecular Organization ◽  
Homologous Chromosomes ◽  
Meiotic Chromosomes ◽  
C Terminus ◽  
Stochastic Optical Reconstruction Microscopy ◽  
Optical Reconstruction ◽  
20 Nm ◽  
Chromosome Axis

During prophase I of meiosis, chromosomes become organized as loop arrays around the proteinaceous chromosome axis. As homologous chromosomes physically pair and recombine, the chromosome axis is integrated into the tripartite synaptonemal complex (SC) as this structure’s lateral elements (LEs). While the components of the mammalian chromosome axis/LE—including meiosis-specific cohesin complexes, the axial element proteins SYCP3 and SYCP2, and the HORMA domain proteins HORMAD1 and HORMAD2—are known, the molecular organization of these components within the axis is poorly understood. Here, using expansion microscopy coupled with 2-color stochastic optical reconstruction microscopy (STORM) imaging (ExSTORM), we address these issues in mouse spermatocytes at a resolution of 10 to 20 nm. Our data show that SYCP3 and the SYCP2 C terminus, which are known to form filaments in vitro, form a compact core around which cohesin complexes, HORMADs, and the N terminus of SYCP2 are arrayed. Overall, our study provides a detailed structural view of the meiotic chromosome axis, a key organizational and regulatory component of meiotic chromosomes.

scholarly journals Microbial Photoinactivation by Visible Light Results in Limited Loss of Membrane Integrity

Antibiotics ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 341
Author(s):  
Katharina Hoenes ◽  
Richard Bauer ◽  
Barbara Spellerberg ◽  
Martin Hessling
Keyword(s):  
Visible Light ◽  
Pseudomonas Fluorescens ◽  
Bacterial Cell ◽  
Bacterial Species ◽  
Membrane Integrity ◽  
Cell Lysis ◽  
Microbial Inactivation ◽  
Bacterial Cells ◽  
Resistance Development ◽  
Transmission Electron

Interest in visible light irradiation as a microbial inactivation method has widely increased due to multiple possible applications. Resistance development is considered unlikely, because of the multi-target mechanism, based on the induction of reactive oxygen species by wavelength specific photosensitizers. However, the affected targets are still not completely identified. We investigated membrane integrity with the fluorescence staining kit LIVE/DEAD® BacLight™ on a Gram positive and a Gram negative bacterial species, irradiating Staphylococcus carnosus and Pseudomonas fluorescens with 405 nm and 450 nm. To exclude the generation of viable but nonculturable (VBNC) bacterial cells, we applied an ATP test, measuring the loss of vitality. Pronounced uptake of propidium iodide was only observed in Pseudomonas fluorescens at 405 nm. Transmission electron micrographs revealed no obvious differences between irradiated samples and controls, especially no indication of an increased bacterial cell lysis could be observed. Based on our results and previous literature, we suggest that visible light photoinactivation does not lead to rapid bacterial cell lysis or disruption. However, functional loss of membrane integrity due to depolarization or inactivation of membrane proteins may occur. Decomposition of the bacterial envelope following cell death might be responsible for observations of intracellular component leakage.

scholarly journals Mapping Neuronal Connectivity Using Stochastic Optical Reconstruction Microscopy (Storm): The Brainstorm Project

2010 ◽  
Vol 98 (3) ◽  
pp. 214a
Author(s):  
Melike Lakadamyali ◽  
Mark Bates ◽  
Hazen Babcock ◽  
Jeff Lichtman ◽  
Xiaowei Zhuang
Keyword(s):  
Neuronal Connectivity ◽  
Stochastic Optical Reconstruction Microscopy ◽  
Optical Reconstruction

scholarly journals Quenched Stochastic Optical Reconstruction Microscopy (qSTORM) with Graphene Oxide

2018 ◽  
Vol 8 (1) ◽  
Author(s):  
Ruiheng Li ◽  
Pantelis Georgiades ◽  
Henry Cox ◽  
Sorasak Phanphak ◽  
Ian S. Roberts ◽  
...  
Keyword(s):  
Graphene Oxide ◽  
Stochastic Optical Reconstruction Microscopy ◽  
Optical Reconstruction

Preparation of Titanium Oxide Nanotube by Hydrothermal Process

2007 ◽  
Vol 124-126 ◽  
pp. 1165-1168 ◽  
Author(s):  
M. Qamar ◽  
Cho Rong Yoon ◽  
Hyo Jin Oh ◽  
Anna Czoska ◽  
K. Park ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Titanium Oxide ◽  
Sol Gel ◽  
Hydrothermal Process ◽  
Hollow Fibers ◽  
Systematic Analysis ◽  
Transmission Electron ◽  
Tio2 Sol ◽  
Naoh Solution

The TiO2 sol was prepared hydrothermally in an autoclave from aqueous TiOCl2 solutions as starting precursor. Hollow fibers were obtained when sol-gel derived TiO2 sol was treated chemically with NaOH solution and subsequently heated in autoclave under various conditions. A systematic analysis of the influence of different NaOH concentrations on the formation of nanotubes has been carried out using XRD and SEM. The phase structure of the synthesized material was determined by transmission electron microscopy and found that these materials are, infact, hollow fibers widely known as nanotubes. From the TEM images, the outer and inner diameters of the tubes were measured ca. 8 and about 4 nm, respectively, with several hundred nanometers in length.

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