scholarly journals Dark-field microscopy enhance visibility of CD31 endothelial staining

2020 ◽  
Vol 64 (3) ◽  
Author(s):  
Eva Jennische ◽  
Stefan Lange ◽  
Ragnar Hultborn
Keyword(s):  
Phase Contrast ◽  
Scattered Light ◽  
Dark Field ◽  
Light Scatter ◽  
Endothelial Lining ◽  
Microscopy Technique ◽  
Dark Field Microscopy ◽  
Dark Field Illumination ◽  
Normal Human ◽  
Intense Light

A simple dark field microscopy technique was used for visualization of blood vessels in normal human renal tissues and carcinoma. Phase contrast condenser ring apt for high power objectives was combined with a 10x objective in order to create a dark field illumination of the specimens examined. The endothelial lining of the vessels had been stained by using CD31 monoclonal antibodies combined with conventional peroxidase immunohistochemistry. The final DAB addition used for this technique induced an intense light scatter in the dark field microscope. This scattered light originating from the endothelial lining made the walls of the bright vessels easily detectable from the dark background.

Dark-field microscopy for characterization of single molecule dynamicsin vitroandin vivo

2019 ◽  
Vol 11 (21) ◽  
pp. 2778-2784
Author(s):  
Bingquan Wang ◽  
Dan Sun ◽  
Ce Zhang ◽  
Kaige Wang ◽  
Jintao Bai
Keyword(s):  
Single Molecule ◽  
Scattered Light ◽  
Dark Field ◽  
Fluorescent Labeling ◽  
Single Molecule Detection ◽  
Dark Field Microscopy

Dark-field microscopy directly detects scattered light from a sample, and therefore requires no fluorescent labeling for single molecule detection.

scholarly journals Genetic manipulation of the optical refractive index in living cells

2020 ◽  
Author(s):  
Junko Ogawa ◽  
Yoko Iwata ◽  
Nina U Tonnu ◽  
Chitra Gopinath ◽  
Ling Huang ◽  
...  
Keyword(s):  
Refractive Index ◽  
Phase Contrast ◽  
Genetic Manipulation ◽  
Physical Property ◽  
Living Cells ◽  
Dark Field ◽  
Dark Field Microscopy ◽  
Protein Nanoparticles ◽  
Cellular Compartments ◽  
Cellular Components

AbstractThe optical refractive index of cellular components is generally not a property considered amenable to manipulation in microscopy as this is an intrinsic physical property of materials. Here we show that by targeting cephalopod reflectin protein nanoparticles one can manipulate the optical refractive index of mammalian cellular compartments. We further demonstrate that refractive index alteration based contrast agents can be utilized for dark field microscopy and quantitative phase contrast holotomography. Additionally we have molecularly cloned novel reflectins with improved and novel optical properties.

Light-microscopic observations of individual microtubules reconstituted from brain tubulin

1975 ◽  
Vol 19 (3) ◽  
pp. 607-620
Author(s):  
R. Kuriyama ◽  
T. Miki-Noumura
Keyword(s):  
Phase Contrast ◽  
Time Course ◽  
Length Distribution ◽  
Dark Field ◽  
Optical Systems ◽  
Viscosity Change ◽  
Dark Field Illumination ◽  
Brain Microtubules ◽  
Brain Tubulin ◽  
Plateau Level

The course of polymerization of individual brain microtubules could be observed with a light microscope employing dark-field illumination. Statistical analysis of the increase in microtubule length during the polymerization was in accordance with the time course of viscosity change of the tubulin solution. After a plateau level in viscosity was attained, there was no significant change in histograms showing length distribution. These observations were confirmed with fixed and stained microtubules, using a phase-contrast microscope. Observations with dark-field illumination revealed that reconstituted microtubules depolymerized and disappeared immediately upon exposure to buffer containing CaCl2 or sulphydryl reagents such as p-chloromercuriphenyl sulphonic acid (PCMPS) and p-chloromercuribenzoic acid (PCMB). They were also cold-labile. The growth of heterogeneous microtubules which were assembled by mixing purified tubulin dimers with ciliary outer fibres could also be followed with these optical systems.

Viewing the Specimen Using Reflected-Light Microscopy

2010 ◽  
pp. 89-114
Keyword(s):  
Composite Materials ◽  
Light Microscopy ◽  
Optical Microscopy ◽  
Polarized Light ◽  
Dark Field ◽  
Microscopy Technique ◽  
Reflected Light ◽  
Specific Analysis ◽  
Dark Field Illumination ◽  
Preparation Techniques

Abstract The analysis of composite materials using optical microscopy is a process that can be made easy and efficient with only a few contrast methods and preparation techniques. This chapter is intended to provide information that will help an investigator select the appropriate microscopy technique for the specific analysis objectives with a given composite material. The chapter opens with a discussion of macrophotography and microscope alignment, and then goes on to describe various illumination techniques that are useful for specific analysis requirements. These techniques include bright-field illumination, dark-field illumination, polarized-light microscopy, interference and contrast microscopy, and fluorescence microscopy. The chapter also provides a discussion of sample preparation materials such as dyes, etchants, and stains for the analysis of composite materials using optical microscopy.

scholarly journals The Fate of Inhaled Nanoparticles: Detection and Measurement by Enhanced Dark-field Microscopy

2017 ◽  
Vol 46 (1) ◽  
pp. 28-46 ◽  
Author(s):  
Robert R. Mercer ◽  
James F. Scabilloni ◽  
Liying Wang ◽  
Lori A. Battelli ◽  
James M. Antonini ◽  
...  
Keyword(s):  
Light Microscope ◽  
High Efficiency ◽  
Scattered Light ◽  
Dark Field ◽  
Potential Health ◽  
Nanoparticle Deposition ◽  
Nanoparticle Distribution ◽  
Dark Field Microscopy ◽  
Microscopic Evaluation ◽  
Inhaled Nanoparticles

Assessing the potential health risks for newly developed nanoparticles poses a significant challenge. Nanometer-sized particles are not generally detectable with the light microscope. Electron microscopy typically requires high-level doses, above the physiologic range, for particle examination in tissues. Enhanced dark-field microscopy (EDM) is an adaption of the light microscope that images scattered light. Nanoparticles scatter light with high efficiency while normal tissues do not. EDM has the potential to identify the critical target sites for nanoparticle deposition and injury in the lungs and other organs. This study describes the methods for EDM imaging of nanoparticles and applications. Examples of EDM application include measurement of deposition and clearance patterns. Imaging of a wide variety of nanoparticles demonstrated frequent situations where nanoparticles detected by EDM were not visible by light microscopy. EDM examination of colloidal gold nanospheres (10–100 nm diameter) demonstrated a detection size limit of approximately 15 nm in tissue sections. EDM determined nanoparticle volume density was directly proportional to total lung burden of exposed animals. The results confirm that EDM can determine nanoparticle distribution, clearance, transport to lymph nodes, and accumulation in extrapulmonary organs. Thus, EDM substantially improves the qualitative and quantitative microscopic evaluation of inhaled nanoparticles.

Elastic Scattering in Electron Microscopy

1973 ◽  
Vol 31 ◽  
pp. 252-253
Author(s):  
J. Langmore ◽  
M. Isaacson ◽  
J. Wall ◽  
A. V. Crewe
Keyword(s):  
Electron Microscopy ◽  
Elastic Scattering ◽  
Cross Section ◽  
Quantum Noise ◽  
Dark Field ◽  
Elastic Cross Section ◽  
Biological Molecules ◽  
Dark Field Microscopy ◽  
Field Systems ◽  
Effects Of Radiation

High resolution dark field microscopy is becoming an important tool for the investigation of unstained and specifically stained biological molecules. Of primary consideration to the microscopist is the interpretation of image Intensities and the effects of radiation damage to the specimen. Ignoring inelastic scattering, the image intensity is directly related to the collected elastic scattering cross section, σɳ, which is the product of the total elastic cross section, σ and the eficiency of the microscope system at imaging these electrons, η. The number of potentially bond damaging events resulting from the beam exposure required to reduce the effect of quantum noise in the image to a given level is proportional to 1/η. We wish to compare η in three dark field systems.

Simulated Dark-Field Electron Micrographs of Point Defects and Organometallics

1975 ◽  
Vol 33 ◽  
pp. 200-201
Author(s):  
William Krakow
Keyword(s):  
Electron Micrographs ◽  
Point Defects ◽  
Structure Determination ◽  
Atomic Structure ◽  
Image Interpretation ◽  
Dark Field ◽  
Field Electron ◽  
Dark Field Microscopy ◽  
Dark Field Electron

Tilted beam dark-field microscopy has been applied to atomic structure determination in perfect crystals, several synthesized molecules with heavy atcm markers and in the study of displaced atoms in crystals. Interpretation of this information in terms of atom positions and atom correlations is not straightforward. Therefore, calculated dark-field images can be an invaluable aid in image interpretation.

Removal of inelastically scattered electrons substantially increases phase contrast on frozen-hydrated molecules

1987 ◽  
Vol 45 ◽  
pp. 652-653
Author(s):  
John P. Langmore ◽  
Brian D. Athey
Keyword(s):  
Electron Diffraction ◽  
Phase Contrast ◽  
Low Dose ◽  
Dark Field ◽  
Biological Structure ◽  
Bright Field ◽  
Low Contrast ◽  
Negative Stain ◽  
The Difference ◽  
Better Than

Although electron diffraction indicates better than 0.3nm preservation of biological structure in vitreous ice, the imaging of molecules in ice is limited by low contrast. Thus, low-dose images of frozen-hydrated molecules have significantly more noise than images of air-dried or negatively-stained molecules. We have addressed the question of the origins of this loss of contrast. One unavoidable effect is the reduction in scattering contrast between a molecule and the background. In effect, the difference in scattering power between a molecule and its background is 2-5 times less in a layer of ice than in vacuum or negative stain. A second, previously unrecognized, effect is the large, incoherent background of inelastic scattering from the ice. This background reduces both scattering and phase contrast by an additional factor of about 3, as shown in this paper. We have used energy filtration on the Zeiss EM902 in order to eliminate this second effect, and also increase scattering contrast in bright-field and dark-field.

Towards 1-Ångstrom-resolution STEM

1993 ◽  
Vol 51 ◽  
pp. 996-997
Author(s):  
H.S. von Harrach ◽  
D.E. Jesson ◽  
S.J. Pennycook
Keyword(s):  
High Resolution ◽  
Field Emission ◽  
Phase Contrast ◽  
Spherical Aberration ◽  
A Priori ◽  
Dark Field ◽  
Image Resolution ◽  
Objective Lens ◽  
Annular Dark Field ◽  
First Results

Phase contrast TEM has been the leading technique for high resolution imaging of materials for many years, whilst STEM has been the principal method for high-resolution microanalysis. However, it was demonstrated many years ago that low angle dark-field STEM imaging is a priori capable of almost 50% higher point resolution than coherent bright-field imaging (i.e. phase contrast TEM or STEM). This advantage was not exploited until Pennycook developed the high-angle annular dark-field (ADF) technique which can provide an incoherent image showing both high image resolution and atomic number contrast.This paper describes the design and first results of a 300kV field-emission STEM (VG Microscopes HB603U) which has improved ADF STEM image resolution towards the 1 angstrom target. The instrument uses a cold field-emission gun, generating a 300 kV beam of up to 1 μA from an 11-stage accelerator. The beam is focussed on to the specimen by two condensers and a condenser-objective lens with a spherical aberration coefficient of 1.0 mm.

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