Demonstration of pulmonary vascular perfusion by electron and light microscopy

1993 ◽  
Vol 75 (4) ◽  
pp. 1877-1883 ◽  
Author(s):  
M. F. Konig ◽  
J. M. Lucocq ◽  
E. R. Weibel
Keyword(s):  
Electron Microscopy ◽  
Light Microscopy ◽  
Light And Electron Microscopy ◽  
Rabbit Serum ◽  
Capillary Perfusion ◽  
Vascular Perfusion ◽  
Thin Sections ◽  
Gold Particles ◽  
Capillary Recruitment

To estimate the fraction of dense pulmonary capillary network that is perfused under physiological conditions, we developed a new method for the demonstration of in vivo capillary perfusion by light and electron microscopy. Blood plasma was labeled by 8-nm colloidal gold particles coated with rabbit serum albumin. In anesthetized rabbits, 4#x2013;5 ml of this tracer were injected into the right atrium. Two and 15 min later, the circulation was interrupted by a snare around the heart, and the lung was fixed by instillation with glutaraldehyde. Gold particles were found in the plasma space of alveolar capillaries as well as in other organs. A random sample of thin sections studied by electron microscopy revealed that the entire capillary bed of the lung was perfused at least with plasma within 2 min after tracer infusion. Light microscopy of silver-enhanced sections showed areas with different staining intensities but no obviously unperfused capillaries. The concept of capillary recruitment, which would require a significant fraction of capillaries unperfused at rest, may have to be reassessed to consider time factors as well as the two-phase nature of blood; red blood cells and plasma may take different paths.

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).

Modification for Electron Miscroscopy of the Corbett Ammoniacal Silver Stain

1971 ◽  
Vol 29 ◽  
pp. 484-485
Author(s):  
Robert L. Corbett
Keyword(s):  
Electron Microscopy ◽  
Electron Microscope ◽  
Light Microscopy ◽  
Low Power ◽  
Light And Electron Microscopy ◽  
Glass Slide ◽  
Silver Stain ◽  
Thin Sections ◽  
Silver Deposits ◽  
Ammoniacal Silver

The ammoniacal silver stain for light microscopy of plastic embedded sections has been adapted for use in electron microscopy. Since the silver will stain even very thin sections, i.e., silver and gold for light microscopy, and silver deposits are sufficiently electron dense to be seen in the electron microscope, the results are very useful for correlating light and electron microscopy. Compared to the conventional stains for electron microscopy which usually take over one-half hour, the silver procedure can be done in five minutes or less and thus provides a quick look at sections This stain has more contrast, so it is especially good for low power electron microscopy. The ability of the silver to stain very thin sections enables a correlation between light and electron microscopy in three ways. First, thin sections can be stained with silver on a glass slide and compared with immediately adjacent thin sections on grids stained the usual way for electron microscopy.

scholarly journals Localization of immunoreactive tyrosine hydroxylase in the goldfish retina with pre-embedding immunolabeling with one-nanometer colloidal gold particles and gold toning.

1992 ◽  
Vol 40 (10) ◽  
pp. 1465-1470 ◽  
Author(s):  
D W Marshak
Keyword(s):  
Electron Microscopy ◽  
Tyrosine Hydroxylase ◽  
Colloidal Gold ◽  
Light And Electron Microscopy ◽  
Rabbit Antiserum ◽  
Gold Chloride ◽  
Thin Sections ◽  
Gold Particles ◽  
Rabbit Igg ◽  
Goldfish Retina

The goal of this study was to develop an alternative to silver intensification for visualizing small colloidal gold particles by light and electron microscopy. The isolated goldfish retina was labeled with rabbit antiserum to tyrosine hydroxylase and 1-nm colloidal gold-conjugated goat anti-rabbit IgG. The gold particles were enlarged by toning with gold chloride, followed by reduction in oxalic acid. Dopaminergic interplexiform cells were clearly visible by light microscopy and, in lightly-fixed material treated with detergent, they were labeled in their entirety. Labeling was qualitatively similar, although less extensive, in material fixed and processed for electron microscopy. The labeled processes were apparent in ultra-thin sections viewed at low magnification, but the gold-toned particles were not so large that they obscured subcellular structures. The procedure apparently had no deleterious effects on the tissue, since the ultrastructural preservation was comparable to that seen with other pre-embedding immunolabeling methods. The technique was simple, reliable and, since the gold solutions were so dilute, relatively inexpensive.

scholarly journals Dense HRP filling in pre-fixed brain tissue for light and electron microscopy.

1987 ◽  
Vol 35 (10) ◽  
pp. 1127-1136 ◽  
Author(s):  
G H Kageyama ◽  
R L Meyer
Keyword(s):  
Electron Microscopy ◽  
Optic Nerve ◽  
High Resolution ◽  
Light Microscopy ◽  
Glial Cells ◽  
Light And Electron Microscopy ◽  
Polyethylene Tube ◽  
Fixed Tissue ◽  
Cat Brain

The use of neuroanatomical markers in tissues that have been pre-fixed has been virtually ignored, even though this approach could offer certain advantages over in vivo methods, in terms of convenience of application and choice of markers. We have found that HRP can be used on well-fixed brains of cats and goldfish to fill neurons, dendrites, axons, terminals, glial cells, and glial processes for high-resolution light microscopy and electron microscopy. Best results were obtained using brains that were perfusion-fixed with 2.5% depolymerized paraformaldehyde and 1.5% glutaraldehyde. Two methods of HRP application were used: optically guided injections of microliter quantities into various regions of cat brain, and optic nerve fills in goldfish by attaching an HRP-filled polyethylene tube for periods of 1 day to 2 weeks. HRP applied in these ways to pre-fixed tissue was found to fill neurons or glial cells with solid label in the anterograde and retrograde directions.

Two- or three-way observations of the identical cell elements within the same sections by LM, TEM and/or SEM

1978 ◽  
Vol 36 (2) ◽  
pp. 82-83 ◽  
Author(s):  
Nakazo Watari ◽  
Yasuaki Hotta ◽  
Yoshio Mabuchi
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Fine Structure ◽  
Light Microscopy ◽  
Thin Sections ◽  
Transmission Electron ◽  
Identical Cell ◽  
Electron Microscopes ◽  
Scanning Electron

It is very useful if we can observe the identical cell elements within the same sections by light microscopy (LM), transmission electron microscopy (TEM) and/or scanning electron microscopy (SEM) sequentially, because, the cell fine structure can not be indicated by LM, while the color is; on the other hand, the cell fine structure can be very easily observed by EM, although its color properties may not. However, there is one problem in that LM requires thick sections of over 1 μm, while EM needs very thin sections of under 100 nm. Recently, we have developed a new method to observe the same cell elements within the same plastic sections using both light and transmission (conventional or high-voltage) electron microscopes.In this paper, we have developed two new observation methods for the identical cell elements within the same sections, both plastic-embedded and paraffin-embedded, using light microscopy, transmission electron microscopy and/or scanning electron microscopy (Fig. 1).

Ectodesmata as Revealed By Freeze-Etch Preparations of Plant Epidermal Cell Walls

1973 ◽  
Vol 31 ◽  
pp. 468-469
Author(s):  
N.C. Lyon ◽  
W. C. Mueller
Keyword(s):  
Electron Microscopy ◽  
Acetic Acid ◽  
Nitric Acid ◽  
Oxalic Acid ◽  
Light Microscopy ◽  
Epidermal Cell ◽  
Cell Walls ◽  
Plant Viruses ◽  
Plant Leaves ◽  
Thin Sections

Schumacher and Halbsguth first demonstrated ectodesmata as pores or channels in the epidermal cell walls in haustoria of Cuscuta odorata L. by light microscopy in tissues fixed in a sublimate fixative (30% ethyl alcohol, 30 ml:glacial acetic acid, 10 ml: 65% nitric acid, 1 ml: 40% formaldehyde, 5 ml: oxalic acid, 2 g: mecuric chloride to saturation 2-3 g). Other workers have published electron micrographs of structures transversing the outer epidermal cell in thin sections of plant leaves that have been interpreted as ectodesmata. Such structures are evident following treatment with Hg++ or Ag+ salts and are only rarely observed by electron microscopy. If ectodesmata exist without such treatment, and are not artefacts, they would afford natural pathways of entry for applied foliar solutions and plant viruses.

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.

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.

Colloidal gold particles as a new in vivo marker of early acute lung injury

2004 ◽  
Vol 287 (4) ◽  
pp. L867-L878 ◽  
Author(s):  
Kai Heckel ◽  
Rainer Kiefmann ◽  
Martina Dörger ◽  
Mechthild Stoeckelhuber ◽  
Alwin E. Goetz
Keyword(s):  
Electron Microscopy ◽  
Acute Lung Injury ◽  
Gas Exchange ◽  
Lung Injury ◽  
Colloidal Gold ◽  
Microvascular Permeability ◽  
Arterial Oxygen ◽  
Control Group ◽  
Gold Particles

Permeability of the endothelial barrier to large molecules plays a pivotal role in the manifestation of early acute lung injury. We present a novel and sensitive technique that brings microanatomical visualization and quantification of microvascular permeability in line. White New Zealand rabbits were anesthetized and ventilated mechanically. Rabbit serum albumin (RSA) was labeled with colloidal gold particles. We quantified macromolecular leakage of gold-labeled RSA and thickening of the gas exchange distance by electron microscopy, taking into account morphology of microvessels. The control group receiving a saline solution represented a normal gas exchange barrier without extravasation of gold-labeled albumin. Infusion of lipopolysaccharide (LPS) resulted in a significant displacement of gold-labeled albumin into pulmonary cells, the lung interstitium, and even the alveolar space. Correspondingly, intravital fluorescence microscopy and digital image analysis indicated thickening of width of alveolar septa. The findings were accompanied by a deterioration of alveolo-arterial oxygen difference, whereas wet/dry ratio and albumin concentration in the bronchoalveolar lavage fluid failed to detect that early stage of pulmonary edema. Inhibition of the nuclear enzyme poly(ADP-ribose) synthetase by 3-aminobenzamide prevented LPS-induced microvascular injury. To summarize: colloidal gold particles visualized by standard electron microscopy are a new and very sensitive in vivo marker of microvascular permeability in early acute lung injury. This technique enabling detailed microanatomical and quantitative pathophysiological characterization of edema formation can form the basis for evaluating novel treatment strategies against acute lung injury.

scholarly journals Post-embedding colloidal gold immunolocalization of laminin to the lamina rara interna, lamina densa, and lamina rara externa of renal glomerular basement membranes.

1986 ◽  
Vol 34 (7) ◽  
pp. 847-853 ◽  
Author(s):  
D R Abrahamson
Keyword(s):  
Colloidal Gold ◽  
Basement Membranes ◽  
Lamina Densa ◽  
Thin Sections ◽  
Gold Particles ◽  
Rabbit Igg ◽  
Gold Immunolocalization ◽  
Lowicryl K4m

Ultrastructural distribution of laminin within renal glomerular (GBM) and tubular basement membranes (TBM) was investigated using post-embedding immunolocalization with colloidal gold. Rat kidneys were fixed with 4% formaldehyde and embedded at 4 degrees C in Lowicryl K4M medium. Thin sections were then sequentially treated with affinity-purified rabbit anti-laminin IgG and anti-rabbit IgG conjugated to 10 nm diameter colloidal gold. Gold bound specifically to the GBM and TBM with particle densities of 690/micron2 and 731/micron2, respectively. In the GBM, the number of gold particles bound/micron2 of lamina densa greater than lamina rara externa greater than lamina rara interna. Closely similar binding patterns were found when kidneys were fixed with 0.5% glutaraldehyde plus 3% formaldehyde and embedded at 60 degrees C in L.R. White resin, but slightly less gold bound to sections overall than that seen with formaldehyde alone and Lowicryl. Taken together, these results illustrate that anti-laminin IgG, whether applied to fixed sections in vitro or introduced in vivo, bound to the lamina rara interna, lamina densa, and lamina rara externa of the GBM and throughout the TBM.

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