scholarly journals A versatile microfluidic device for highly inclined thin illumination microscopy in the moss Physcomitrella patens

2019 ◽  
Author(s):  
Kozgunova Elena ◽  
Gohta Goshima
Keyword(s):  
High Resolution ◽  
Microfluidic Device ◽  
Membrane Trafficking ◽  
Cell Walls ◽  
Physcomitrella Patens ◽  
Plant Cells ◽  
Microscopy Imaging ◽  
Cellular Processes ◽  
High Resolution Microscopy

AbstractHigh-resolution microscopy is a valuable tool to study cellular processes, such as signalling, membrane trafficking, or cytoskeleton remodelling. Several techniques of inclined illumination microscopy allow imaging at near single molecular level; however, the application of these methods to plant cells is limited, due to thick cell walls and necessity to excise a part of the tissue for sample preparation. In this study, we developed simple, easy-to-use microfluidic device for highly inclined and laminated optical sheet (HILO) microscopy using a model plant Physcomitrella patens. We demonstrated that microfluidic device can be used to culture living cells and enables high-resolution HILO imaging of microtubules without perturbing their dynamics. In addition, our microdevice enables the supply and robust washout of compounds during HILO microscopy imaging, for example to perform microtubule regrowth assay. Furthermore, we tested long-term (48 h) HILO imaging using a microdevice and visualised the developmental changes in the microtubule dynamics during tissue regeneration. The microfluidic device designed in this study provides a novel tool to conduct long-term HILO microscopy and washout assays using plant cells.

scholarly journals A versatile microfluidic device for highly inclined thin illumination microscopy in the moss Physcomitrella patens

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Elena Kozgunova ◽  
Gohta Goshima
Keyword(s):  
High Resolution ◽  
Microfluidic Device ◽  
Membrane Trafficking ◽  
Physcomitrella Patens ◽  
Plant Cells ◽  
Microscopy Imaging ◽  
Living Plant ◽  
Cellular Processes ◽  
Novel Applications

Abstract High-resolution microscopy is a valuable tool for studying cellular processes, such as signalling, membrane trafficking, or cytoskeleton remodelling. Several techniques of inclined illumination microscopy allow imaging at a near single molecular level; however, the application of these methods to plant cells is limited, owing to thick cell walls as well as the necessity to excise a part of the tissue for sample preparation. In this study, we utilised a simple, easy-to-use microfluidic device for highly inclined and laminated optical sheet (HILO) microscopy using a model plant Physcomitrella patens. We demonstrated that the shallow microfluidic device can be used for long-term culture of living cells and enables high-resolution HILO imaging of microtubules without perturbing their dynamics. In addition, our microdevice allows the supply and robust washout of compounds during HILO microscopy imaging, for example, to perform a microtubule regrowth assay. Furthermore, we tested long-term (48 h) HILO imaging using a microdevice and visualised the developmental changes in the microtubule dynamics during tissue regeneration. These novel applications of the microfluidic device provide a valuable resource for studying molecular dynamics in living plant cells.

Drink or drive: competition between macropinocytosis and cell migration

2015 ◽  
Vol 43 (1) ◽  
pp. 129-132 ◽  
Author(s):  
Douwe M. Veltman
Keyword(s):  
Cell Migration ◽  
High Resolution ◽  
Cell Motility ◽  
Actin Filaments ◽  
Cellular Processes ◽  
Heavy Drinkers ◽  
Molecular Components ◽  
High Resolution Microscopy ◽  
Wave Complex

The cytoskeleton is utilized for a variety of cellular processes, including migration, endocytosis and adhesion. The required molecular components are often shared between different processes, but it is not well understood how the cells balance their use. We find that macropinocytosis and cell migration are negatively correlated. Heavy drinkers move only slowly and vice versa, fast cells do not take big gulps. Both processes are balanced by the lipid phosphatidylinositol 3,4,5-trisphosphate (PIP3). Elevated PIP3 signalling causes a shift towards macropinocytosis and inhibits motility by redirecting the SCAR/WAVE complex, a major nucleator of actin filaments. High resolution microscopy shows that patches with high levels of PIP3 recruit SCAR/WAVE on their periphery, resulting in circular ruffle formation and engulfment. Results shed new light on the role of PIP3, which is commonly thought to promote cell motility.

scholarly journals Differential Polarization Imaging of Plant Cells. Mapping the Anisotropy of Cell Walls and Chloroplasts

2021 ◽  
Vol 22 (14) ◽  
pp. 7661
Author(s):  
Jasna Simonović Radosavljević ◽  
Aleksandra Lj. Mitrović ◽  
Ksenija Radotić ◽  
László Zimányi ◽  
Győző Garab ◽  
...  
Keyword(s):  
Light Microscopy ◽  
Plant Cell ◽  
Cell Walls ◽  
Laser Scanning ◽  
Imaging Techniques ◽  
Plant Cells ◽  
Quantitative Information ◽  
Laser Scanning Microscopy ◽  
Molecular Organization ◽  
Microscopy Imaging

Modern light microscopy imaging techniques have substantially advanced our knowledge about the ultrastructure of plant cells and their organelles. Laser-scanning microscopy and digital light microscopy imaging techniques, in general—in addition to their high sensitivity, fast data acquisition, and great versatility of 2D–4D image analyses—also opened the technical possibilities to combine microscopy imaging with spectroscopic measurements. In this review, we focus our attention on differential polarization (DP) imaging techniques and on their applications on plant cell walls and chloroplasts, and show how these techniques provided unique and quantitative information on the anisotropic molecular organization of plant cell constituents: (i) We briefly describe how laser-scanning microscopes (LSMs) and the enhanced-resolution Re-scan Confocal Microscope (RCM of Confocal.nl Ltd. Amsterdam, Netherlands) can be equipped with DP attachments—making them capable of measuring different polarization spectroscopy parameters, parallel with the ‘conventional’ intensity imaging. (ii) We show examples of different faces of the strong anisotropic molecular organization of chloroplast thylakoid membranes. (iii) We illustrate the use of DP imaging of cell walls from a variety of wood samples and demonstrate the use of quantitative analysis. (iv) Finally, we outline the perspectives of further technical developments of micro-spectropolarimetry imaging and its use in plant cell studies.

High Resolution Microscopy of Multi-Layered Biological Crystals

1982 ◽  
Vol 40 ◽  
pp. 80-83
Author(s):  
H.A. Cohen ◽  
T.W. Jeng ◽  
W. Chiu
Keyword(s):  
High Resolution ◽  
Low Dose ◽  
Three Dimensional ◽  
Data Interpretation ◽  
Two Dimensional ◽  
Biological Objects ◽  
Density Maps ◽  
Density Map ◽  
Complex Crystal ◽  
High Resolution Microscopy

This tutorial will discuss the methodology of low dose electron diffraction and imaging of crystalline biological objects, the problems of data interpretation for two-dimensional projected density maps of glucose embedded protein crystals, the factors to be considered in combining tilt data from three-dimensional crystals, and finally, the prospects of achieving a high resolution three-dimensional density map of a biological crystal. This methodology will be illustrated using two proteins under investigation in our laboratory, the T4 DNA helix destabilizing protein gp32*I and the crotoxin complex crystal.

High Resolution Observations of Ion-Milling Induced Transformation in Synthetic Hollandses

1981 ◽  
Vol 39 ◽  
pp. 114-115 ◽  
Author(s):  
T. J. Headley
Keyword(s):  
High Resolution ◽  
Radioactive Waste Disposal ◽  
Ion Milling ◽  
Minor Elements ◽  
Waste Forms ◽  
Carbon Substrates ◽  
Structural Differences ◽  
Oxide Phases ◽  
Argon Ions ◽  
High Resolution Microscopy

Oxide phases having the hollandite structure have been identified in multiphase ceramic waste forms being developed for radioactive waste disposal. High resolution studies of phases in the waste forms described in Ref. [2] were initiated to examine them for fine scale structural differences compared to natural mineral analogs. Two hollandites were studied: a (Ba,Cs,K)-titan-ate with minor elements in solution that is produced in the waste forms, and a synthesized BaAl2Ti6O16 phase containing ∼ 4.7 wt% Cs2O. Both materials were consolidated by hot pressing at temperatures above 1100°C. Samples for high resolution microscopy were prepared both by ion-milling (7kV argon ions) and by crushing and dispersing the fragments on holey carbon substrates. The high resolution studies were performed in a JEM 200CX/SEG operating at 200kV.

Design of a new objective lens for an HB-501A STEM

1991 ◽  
Vol 49 ◽  
pp. 416-417
Author(s):  
Earl J. Kirkland ◽  
Robert J. Keyse
Keyword(s):  
High Resolution ◽  
Spherical Aberration ◽  
Scanning Transmission Electron Microscope ◽  
Dark Field ◽  
Objective Lens ◽  
Specimen Holder ◽  
Transmission Electron ◽  
Scanning Transmission ◽  
Annular Dark Field ◽  
High Resolution Microscopy

An ultra-high resolution pole piece with a coefficient of spherical aberration Cs=0.7mm. was previously designed for a Vacuum Generators HB-501A Scanning Transmission Electron Microscope (STEM). This lens was used to produce bright field (BF) and annular dark field (ADF) images of (111) silicon with a lattice spacing of 1.92 Å. In this microscope the specimen must be loaded into the lens through the top bore (or exit bore, electrons traveling from the bottom to the top). Thus the top bore must be rather large to accommodate the specimen holder. Unfortunately, a large bore is not ideal for producing low aberrations. The old lens was thus highly asymmetrical, with an upper bore of 8.0mm. Even with this large upper bore it has not been possible to produce a tilting stage, which hampers high resolution microscopy.

Practical High Resolution Microscopy

1968 ◽  
Vol 26 ◽  
pp. 302-303
Author(s):  
P. A. Marsh ◽  
T. Mullens ◽  
D. Price
Keyword(s):  
High Resolution ◽  
Magnetic Fields ◽  
Low Frequency ◽  
Resolving Power ◽  
Careful Attention ◽  
Electron Microscopes ◽  
The Relationship ◽  
High Resolution Microscopy ◽  
New Facilities

It is possible to exceed the guaranteed resolution on most electron microscopes by careful attention to microscope parameters essential for high resolution work. While our experience is related to a Philips EM-200, we hope that some of these comments will apply to all electron microscopes.The first considerations are vibration and magnetic fields. These are usually measured at the pre-installation survey and must be within specifications. It has been our experience, however, that these factors can be greatly influenced by the new facilities and therefore must be rechecked after the installation is completed. The relationship between the resolving power of an EM-200 and the maximum tolerable low frequency interference fields in milli-Oerstedt is 10 Å - 1.9, 8 Å - 1.4, 6 Å - 0.8.

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