scholarly journals Fluorescent and Electron-Dense Green Color Emitting Nanodiamonds for Single-Cell Correlative Microscopy

Molecules ◽  
2020 ◽  
Vol 25 (24) ◽  
pp. 5897 ◽  
Author(s):  
Neeraj Prabhakar ◽  
Markus Peurla ◽  
Olga Shenderova ◽  
Jessica M. Rosenholm
Keyword(s):  
Electron Microscopy ◽  
Electron Beam ◽  
Light Microscopy ◽  
Light And Electron Microscopy ◽  
Fluorescent Properties ◽  
Green Color ◽  
Chemical Treatments ◽  
A Cell ◽  
Green Fluorescent ◽  
Microscopy Images

Correlative light and electron microscopy (CLEM) is revolutionizing how cell samples are studied. CLEM provides a combination of the molecular and ultrastructural information about a cell. For the execution of CLEM experiments, multimodal fiducial landmarks are applied to precisely overlay light and electron microscopy images. Currently applied fiducials such as quantum dots and organic dye-labeled nanoparticles can be irreversibly quenched by electron beam exposure during electron microscopy. Generally, the sample is therefore investigated with a light microscope first and later with an electron microscope. A versatile fiducial landmark should offer to switch back from electron microscopy to light microscopy while preserving its fluorescent properties. Here, we evaluated green fluorescent and electron dense nanodiamonds for the execution of CLEM experiments and precisely correlated light microscopy and electron microscopy images. We demonstrated that green color emitting fluorescent nanodiamonds withstand electron beam exposure, harsh chemical treatments, heavy metal straining, and, importantly, their fluorescent properties remained intact for light microscopy.

The Response of Testis Cells to Low Dose X-Irradiation

1981 ◽  
Vol 39 ◽  
pp. 542-543
Author(s):  
D. E. Philpott ◽  
W. Sapp ◽  
C. Williams ◽  
Joann Stevenson ◽  
S. Black
Keyword(s):  
Electron Microscopy ◽  
Stem Cell ◽  
Light Microscopy ◽  
Dose Level ◽  
Cross Sections ◽  
Low Dose ◽  
Light And Electron Microscopy ◽  
Cellular Responses ◽  
Post Irradiation ◽  
X Irradiation

The response of spermatogonial cells to X-irradiation is well documented. It has been shown that there is a radiation resistent stem cell (As) which, after irradiation, replenishes the seminiferous epithelium. Most investigations in this area have dealt with radiation dosages of 100R or more. This study was undertaken to observe cellular responses at doses less than 100R of X-irradiation utilizing a system in which the tissue can be used for light and electron microscopy.Brown B6D2F1 mice aged 16 weeks were exposed to X-irradiation (225KeV; 15mA; filter 0.35 Cu; 50-60 R/min). Four mice were irradiated at each dose level between 1 and 100 rads. Testes were removed 3 days post-irradiation, fixed, and embedded. Sections were cut at 2 microns for light microscopy. After staining, surviving spermatogonia were identified and counted in tubule cross sections. The surviving fraction of spermatogonia compared to control, S/S0, was plotted against dose to give the curve shown in Fig. 1.

Artifacts caused by dehydration and epoxy embedding in TEM

1991 ◽  
Vol 49 ◽  
pp. 338-339
Author(s):  
Hilton H. Mollenhauer
Keyword(s):  
Electron Microscopy ◽  
Electron Beam ◽  
Volume Change ◽  
Tissue Damage ◽  
Beam Specimen ◽  
Chemical Fixation ◽  
Resin Impregnation ◽  
Storage Vesicles ◽  
A Cell ◽  
Tissue Shrinkage

Various means have been devised to preserve biological specimens for electron microscopy, the most common being chemical fixation followed by dehydration and resin impregnation. It is intuitive, and has been amply demonstrated, that these manipulations lead to aberrations of many tissue elements. This report deals with three parts of this problem: specimen dehydration, epoxy embedding resins, and electron beam-specimen interactions. However, because of limited space, only a few points can be summarized.Dehydration: Tissue damage, or at least some molecular transitions within the tissue, must occur during passage of a cell or tissue to a nonaqueous state. Most obvious, perhaps, is a loss of lipid, both that which is in the form of storage vesicles and that associated with tissue elements, particularly membranes. Loss of water during dehydration may also lead to tissue shrinkage of 5-70% (volume change) depending on the tissue and dehydrating agent.

Solving the mechanisms responsible for organelle behavior in vivo by same-cell correlative light and electron microscopy

1992 ◽  
Vol 50 (1) ◽  
pp. 14-15
Author(s):  
Conly L. Rieder
Keyword(s):  
Electron Microscopy ◽  
Quantitative Analysis ◽  
Light Microscopy ◽  
Living Cell ◽  
Light And Electron Microscopy ◽  
Time Behavior ◽  
Dynamic Interactions ◽  
Positive Identification ◽  
Cellular Components

The behavior of many cellular components, and their dynamic interactions, can be characterized in the living cell with considerable spatial and temporal resolution by video-enhanced light microscopy (video-LM). Indeed, under the appropriate conditions video-LM can be used to determine the real-time behavior of organelles ≤ 25-nm in diameter (e.g., individual microtubules—see). However, when pushed to its limit the structures and components observed within the cell by video-LM cannot be resolved nor necessarily even identified, only detected. Positive identification and a quantitative analysis often requires the corresponding electron microcopy (EM).

Development of an in vitro Model for the Study of the Response of Equine Tendon Fibroblasts to Injury and Medication

1997 ◽  
Vol 10 (01) ◽  
pp. 6-11 ◽  
Author(s):  
R. F. Rosenbusch ◽  
L. C. Booth ◽  
L. A. Dahlgren
Keyword(s):  
Electron Microscopy ◽  
Extracellular Matrix ◽  
Light Microscopy ◽  
Light And Electron Microscopy ◽  
Tendon Healing ◽  
Matrix Proteins ◽  
Isolated Cells ◽  
Superficial Digital Flexor Tendon ◽  
Vitro Model

SummaryEquine tendon fibroblasts were isolated from explants of superficial digital flexor tendon, subcultured and maintained in monolayers. The cells were characterized by light microscopy, electron microscopy and radiolabel studies for proteoglycan production. Two predominant cell morphologies were identified. The cells dedifferentiated toward a more spindle shape with repeated subcultures. Equine tendon fibroblasts were successfully cryopreserved and subsequently subcultured. The ability to produce proteoglycan was preserved.The isolated cells were identified as fibroblasts, based on their characteristic shape by light microscopy and ultrastructure and the active production of extracellular matrix proteins. Abundant rough endoplasmic reticulum and the production of extracellular matrix products demonstrated active protein production and export. Proteoglycans were measurable via liquid scintillation counting in both the cell-associated fraction and free in the supernatant. This model is currently being utilized to study the effects of polysulfated glycosaminoglycan on tendon healing. Future uses include studying the effects of other pharmaceuticals, such as hyaluronic acid, on tendon healing.A model was developed for in vitro investigations into tendon healing. Fibroblasts were isolated from equine superficial digital flexor tendons and maintained in monolayer culture. The tenocytes were characterized via light and electron microscopy. Proteoglycan production was measured, using radio-label techniques. The fibroblasts were cryopreserved and subsequently subcultured. The cells maintained their capacity for proteoglycan production, following repeated subculturing and cryopreservation.

scholarly journals FINE STRUCTURAL OBSERVATIONS RELATING TO THE PRODUCTION OF COLOR BY THE IRIDOPHORES OF A LIZARD, ANOLIS CAROLINENSIS

1972 ◽  
Vol 53 (1) ◽  
pp. 38-52 ◽  
Author(s):  
Susannah T. Rohrlich ◽  
Keith R. Porter
Keyword(s):  
Thin Film ◽  
Electron Microscopy ◽  
Light And Electron Microscopy ◽  
Structural Data ◽  
Anolis Carolinensis ◽  
Dorsal Skin ◽  
Green Color ◽  
Structural Details ◽  
Thin Film Interference ◽  
Crystal Arrangement

This paper presents the results of light and electron microscopy done on iridophores in the dorsal skin of the lizard Anolis carolinensis. New fine-structural details are revealed, and their importance is discussed. Of some interest is the complex of filaments between crystalline sheets in the cell. It is proposed that this complex is involved in the arrangement of crystals into crystalline sheets, and that the crystal arrangement and spacing are critical for the production of the cells' blue-green color. Tyndall scattering and thin-film interference are discussed as possible explanations for iridophore color production in relation to the fine-structural data obtained.

scholarly journals An Easy Path for Correlative Electron and Super-Resolution Light Microscopy

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Dorothea Pinotsi ◽  
Simona Rodighiero ◽  
Silvia Campioni ◽  
Gabor Csucs
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Light Microscopy ◽  
Amyloid Fibrils ◽  
Light And Electron Microscopy ◽  
Super Resolution ◽  
Wide Field ◽  
Set Up ◽  
Bio Compatibility ◽  
Scanning Electron

Abstract A number of new Correlative Light and Electron Microscopy approaches have been developed over the past years, offering the opportunity to combine the specificity and bio-compatibility of light microscopy with the high resolution achieved in electron microscopy. More recently, these approaches have taken one step further and also super-resolution light microscopy was combined with transmission or scanning electron microscopy. This combination usually requires moving the specimen between different imaging systems, an expensive set-up and relatively complicated imaging workflows. Here we present a way to overcome these difficulties by exploiting a commercially available wide-field fluorescence microscope integrated in the specimen chamber of a Scanning Electron Microscope (SEM) to perform correlative LM/EM studies. Super-resolution light microscopy was achieved by using a recently developed algorithm - the Super-Resolution Radial Fluctuations (SRRF) - to improve the resolution of diffraction limited fluorescent images. With this combination of hardware/software it is possible to obtain correlative super-resolution light and scanning electron microscopy images in an easy and fast way. The imaging workflow is described and demonstrated on fluorescently labelled amyloid fibrils, fibrillar protein aggregates linked to the onset of multiple neurodegenerative diseases, revealing information about their polymorphism.

scholarly journals THE GLOMERULUS IN EXPERIMENTAL RENAL DISEASE IN RATS AS OBSERVED BY LIGHT AND ELECTRON MICROSCOPY

1955 ◽  
Vol 102 (5) ◽  
pp. 573-580 ◽  
Author(s):  
Carolyn F. Piel ◽  
Luther Dong ◽  
F.W.S. Modern ◽  
Joseph R. Goodman ◽  
Roger Moore
Keyword(s):  
Electron Microscopy ◽  
Endothelial Cells ◽  
Basement Membrane ◽  
Light Microscopy ◽  
Light And Electron Microscopy ◽  
Electron Microscopic ◽  
Crescent Formation ◽  
Colloidal Iron ◽  
Narrow Channels ◽  
Glomerular Capillary

Nephrotoxic serum disease in rats has been studied by light and electron microscopy from 1 hour to 10 weeks after production of the disease. By light microscopy leucocytic infiltration of the glomerular capillary was observed between the 3rd and 6th hour. At 6 hours an increase in colloidal iron-positive material was observed coating the extraluminal surface of the capillaries. Also at this time swelling of the endothelial cells becomes prominent. By 72 hours, thickening of the basement membrane was observed. Glomerular capillary thrombi were observed in approximately half the tissue examined in the first 2 weeks of disease. 50 per cent of the animals showed severe chronic lesions, exudation into the capsular space, crescent formation, and obliteration of glomeruli. At 1 hour electron microscopic pictures showed that osmophilic material may line the foot processes of the epithelial cells and obliterate all but narrow channels of the space between the feet. By 6 hours thickening of the basement membrane was prominent. This change persisted throughout 10 weeks of observation. The tissue from animals which had severe chronic alterations by light microscopy revealed changes which could not be interpreted at this time.

scholarly journals A tribute to Shinya Inoue and innovation in light microscopy

2004 ◽  
Vol 165 (1) ◽  
pp. 21-26 ◽  
Author(s):  
Karen R. Dell ◽  
Ronald D. Vale
Keyword(s):  
Electron Microscopy ◽  
Light Microscopy ◽  
Light Microscope ◽  
Light And Electron Microscopy ◽  
Single Molecules ◽  
Living Cells ◽  
Dynamic Processes ◽  
New Techniques

The 2003 International Prize for Biology was awarded to Shinya Inoue for his pioneering work in visualizing dynamic processes within living cells using the light microscope. He and his scientific descendants are now pushing light microscopy even further by developing new techniques such as imaging single molecules, visualizing processes in living animals, and correlating results from light and electron microscopy.

Scale structure and taxonomy of some species of Raphidocystis, Raphidiophrys, and Pompholyxophrys (Heliozoea) including descriptions of six new taxa

1985 ◽  
Vol 63 (8) ◽  
pp. 1944-1961 ◽  
Author(s):  
K. H. Nicholls ◽  
M. Dürrschmidt
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
New Zealand ◽  
Light Microscopy ◽  
Light And Electron Microscopy ◽  
Scale Structure ◽  
Taxonomic Descriptions ◽  
Transmission Electron ◽  
New Information ◽  
Freshwater Habitats

Sixteen taxa of the genera Raphidocystis, Raphidiophrys, and Pompholyxophrys from freshwater habitats in Canada, Chile, and New Zealand were studied by light and electron microscopy. Six taxa are described as new: Raphidocystis glabra, Raphidiophrys minuta, Raphidiophrys orbicularis ssp. orbicularis, R. orbicularis ssp. ovalis, Pompholyxophrys stellata, and P. ossea. New information on scale structure and arrangement based on scanning and transmission electron microscopy amplifies the taxonomic descriptions of Raphidiophrys ambigua, R. pallida, R. elegans, R. intermedia, R. marginata, R. symmetrica, Pompholyxophrys punicea, P. exigua, and P. ovuligera, which were previously imperfectly known by light microscopy only.

scholarly journals Dynamic-ultrastructural cell volume (3D) correlative microscopy facilitated by intracellular fluorescent nanodiamonds as multi-modal probes

2019 ◽  
Author(s):  
Neeraj Prabhakar ◽  
Ilya Belevich ◽  
Markus Peurla ◽  
Xavier Heiligenstein ◽  
Huan-Cheng Chang ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Cell Volume ◽  
Light And Electron Microscopy ◽  
Three Dimensional ◽  
Correlative Microscopy ◽  
Straightforward Method ◽  
Functional Aspects ◽  
A Cell ◽  
Block Face ◽  
Fluorescent Nanodiamonds

ABSTRACTThree-dimensional correlative light and electron microscopy (3D CLEM) are attaining popularity as a potential technique to explore the functional aspects of a cell together with high-resolution ultrastructural details across the cell volume. In order to perform such a 3D CLEM experiment, there is an imperative requirement for multi-modal probes that are both fluorescent and electron-dense. These multi-modal probes will serve as landmarks in matching up the large full cell volume datasets acquired by different imaging modalities. Fluorescent nanodiamonds (FNDs) are a unique nanosized, fluorescent, and electron-dense material from the nanocarbon family. We hereby propose a novel and straightforward method for executing 3D CLEM using FNDs as multi-modal landmarks. We demonstrate that FNDs is biocompatible and easily identified both in living cell fluorescence imaging and in serial block-face scanning electron microscopy (SB-EM). We illustrate the 3D CLEM method by registering multi-modal datasets.

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