Novel method of simultaneous multiple immunogold localization on resin sections in high resolution scanning electron microscopy

Nebesářová, Jana; Wandrol, Petr; Vancová, Marie

doi:10.1016/j.nano.2015.09.008

Cited by 5 publications

(4 citation statements)

References 9 publications

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“…al. [10] is that it is necessary to constantly switch them. When the accelerating voltage is switched, it causes the electron beam to defocus and shift to a different position.…”

Section: Discussionmentioning

confidence: 99%

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An Advanced Fast Method for the Evaluation of Multiple Immunolabelling Using Gold Nanoparticles Based on Low-Energy Stem

Kitzberger

Yang

Týč

et al. 2023

Preprint

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“…al. [10] is that it is necessary to constantly switch them. When the accelerating voltage is switched, it causes the electron beam to defocus and shift to a different position.…”

Section: Discussionmentioning

confidence: 99%

“…[8. 9] The most recent technique for multiple immunogold labelling was suggested by Nebesarova et al [10] and uses SEM operating in STEM mode. This method is based on distinguishing Au nanoparticles located on the top and bottom of an ultrathin section labelled on both sides.…”

Section: Introductionmentioning

confidence: 99%

An Advanced Fast Method for the Evaluation of Multiple Immunolabelling Using Gold Nanoparticles Based on Low-Energy Stem

Kitzberger

Yang

Týč

et al. 2023

Preprint

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“…Proper vitrification of bulk specimens is another limiting factor for observation in cryo-SEM. Biological cryo-specimens are highly sensitive to electron-beam radiation damage, therefore electron beam exposures must be minimized and observation at very low energy (e.g., 1 kV) enable collection only “topographic” surface information (Nebesářová et al, 2016). …”

Section: Introductionmentioning

confidence: 99%

Pleomorphism and Viability of the Lyme Disease Pathogen Borrelia burgdorferi Exposed to Physiological Stress Conditions: A Correlative Cryo-Fluorescence and Cryo-Scanning Electron Microscopy Study

et al. 2017

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To understand the response of the Lyme disease spirochete Borrelia burgdorferi exposed to stress conditions and assess the viability of this spirochete, we used a correlative cryo-fluorescence and cryo-scanning microscopy approach. This approach enables simple exposition of bacteria to various experimental conditions that can be stopped at certain time intervals by cryo-immobilization, examination of cell viability without necessity to maintain suitable culture conditions during viability assays, and visualization of structures in their native state at high magnification. We focused on rare and transient events e.g., the formation of round bodies and the presence of membranous blebs in spirochetes exposed to culture medium, host sera either without or with the bacteriolytic effect and water. We described all crucial steps of the workflow, particularly the influence of freeze-etching and accelerating voltage on the visualization of topography. With the help of newly designed cryo-transport device, we achieved greater reproducibility.

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“…A higher accelerating voltage enlarge the interaction volume from which the electrons are scattered [ 14 – 16 ]. In ultrathin sections, SEM working in the transmission mode at higher accelerating voltage provide a transmission image of cell structures similarly to TEM [ 16 , 17 ]. At accelerating voltage below 5 kV, SEM provides detailed information on the surface morphology with minimal charging and radiation damage of sensitive (either dried or hydrated) biological samples.…”

Section: Introductionmentioning

confidence: 99%

Correlative Fluorescence and Scanning Electron Microscopy of Labelled Core Fucosylated Glycans Using Cryosections Mounted on Carbon-Patterned Glass Slides

Vancová

Nebesářová

2015

PLoS ONE

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The aim of the study is co-localization of N-glycans with fucose attached to N-acetylglucosamine in α1,3 linkage, that belong to immunogenic carbohydrate epitopes in humans, and N-glycans with α1,6-core fucose typical for mammalian type of N-linked glycosylation. Both glycan epitopes were labelled in cryosections of salivary glands isolated from the tick Ixodes ricinus. Salivary glands secrete during feeding many bioactive molecules and influence both successful feeding and transmission of tick-borne pathogens. For accurate and reliable localization of labelled glycans in both fluorescence and scanning electron microscopes, we used carbon imprints of finder or indexed EM grids on glass slides. We discuss if the topographical images can provide information about labelled structures, the working setting of the field-emission scanning electron microscope and the influence of the detector selection (a below-the-lens Autrata improved YAG detector of back-scattered electrons; in-lens and conventional Everhart-Thornley detectors of secondary electrons) on the imaging of gold nanoparticles, quantum dots and osmium-stained membranes.

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Novel method of simultaneous multiple immunogold localization on resin sections in high resolution scanning electron microscopy

Cited by 5 publications

References 9 publications

An Advanced Fast Method for the Evaluation of Multiple Immunolabelling Using Gold Nanoparticles Based on Low-Energy Stem

An Advanced Fast Method for the Evaluation of Multiple Immunolabelling Using Gold Nanoparticles Based on Low-Energy Stem

Pleomorphism and Viability of the Lyme Disease Pathogen Borrelia burgdorferi Exposed to Physiological Stress Conditions: A Correlative Cryo-Fluorescence and Cryo-Scanning Electron Microscopy Study

Correlative Fluorescence and Scanning Electron Microscopy of Labelled Core Fucosylated Glycans Using Cryosections Mounted on Carbon-Patterned Glass Slides

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