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1.
A method for embedding tissue culture preparations, grown on Millipore filters, in Araldite is described. Accurate light microscopic localization of areas suitable for ultrathin sectioning is possible. The plane of sectioning relative to the Millipore filter can be varied to suit individual experimental needs.  相似文献   

2.
Inner surfaces and fracture faces of rabbit kidney tissue were investigated with high-resolution scanning electron microscopy using two different cryopreparation techniques: (i) for the observation of fracture faces, cryofixed tissue was fractured and coated in a cryopreparation chamber dedicated to SEM, vacuum transferred onto a cold stage and observed in the frozen-hydrated state; (ii) for the observation of inner surfaces of the nephron, water was removed after freezing and fracturing by freeze substitution and critical-point drying of the tissue. By both methods, macromolecular structures such as intramembranous particles on fracture faces and particles on inner surfaces were imaged. The latter method was used to investigate in more detail surface structures of cells in the cortical collecting duct. These studies revealed a heterogeneity of intercalated cells not described thus far.  相似文献   

3.
The crystallography of recrystallization has been investigated in channel‐die deformed pure aluminium bicrystals with {100}<011>/{110}<001> orientations. The microstructural and microtextural changes during the early stages of recrystallization were followed by systematic local orientation measurements using scanning and transmission electron microscopes. In particular, orientation mapping combined with in situ sample heating was used to investigate the formation and growth of new grains at very early stages of recrystallization. Grain boundary migration and ‘consumption’ of the as‐deformed areas was always favoured along directions parallel to the traces of the {111} slip planes that had been most active during deformation.  相似文献   

4.
The maintenance and true pictorial representation of the complex cyto-architecture of tissues is the aim of the preparatory methods for scanning electron microscopy. The procedure employed eliminates complications associated with and due to the vacuum technology necessarily employed in electron optic instruments using a scanning beam. Tissues are fixed by vascular perfusion after the administration of heparin eliminating clot formation. The infusion of Araldite in the tissues prevents the collapse and disorganization of cells due to the combined effects of dehydration, the vacuum of the scanning electron microscope and the scanning beam. The specimens are scoured with acetone and the blocks cured, this latter process ensures their permanence; the standard procedures of gold coating, stub mounting and painting the bases of specimens with a colloidal silver solution are then carried out. The method allows magnifications of the order of 10,000 diameters and an acceptable resolution of some 50 nm is obtained. Cytoarchitectural features demonstrated by the method described are borne out by transmission microscopy.  相似文献   

5.
Using recent papers on scanning electron microscopy (SEM) of chemical vapor deposition (CVD) diamond films, two analytical applications of the SEM are discussed: the morphologic investigations (secondary electron emission mode) and the recognition of impurities and defects [cathodoluminescence (CL) mode]. Studies of CVD diamond films by SEM demonstrate that the morphologies of these films are affected by synthesis conditions, especially by substrate temperature, methane concentration, and total pressure in the reactor. CL spectra and images are useful tools for clarifying the relationship between emission centers and different types of defects generated during the process of diamond crystal growth. The paper shows that the investigations of the morphology, crystallinity, local CL emission, as well as the surface distribution of CL spectra on CVD diamond films by SEM led to the correlative information for quality estimation of films in comparison with natural diamond.  相似文献   

6.
Two simple techniques are described and illustrated. The first is for the study of one specimen by both light microscopy (LM) and scanning electron microscopy (SEM). The second is for the study of one selected specimen by LM, SEM and in ultrathin section by transmission electron microscopy (TEM). Although these techniques were developed for the comparative study of Precambrian organic walled microfossils (OWMs), they could be used for a wide range of other specimens.  相似文献   

7.
Alkali digestion has been used to remove cellular elements of tissues thus exposing the underlying connective tissue framework. We studied the action of this severe alkali treatment on the delicate tissues of rat lung. The lungs of male Sprague-Dawley rats were perfused with saline to remove blood and then inflated by fixation through the airways at 20 cm pressure. Sections of lung 2 × 5 × 5 mm were immersed in 2.5 M NaOH at 25°C for 6 h, 16 h, 24 h, 48 h, and 72 h. The alkali was changed daily. Tissues were washed to neutral with water (24 h), treated with tannic acid (1%, 3h), post-fixed with osmic acid (1%, 3 h) and processed for SEM. At 6 h, epithelial cells started to peel off the alveolar surface. At 16 h the digestion process was well advanced. At 48 h the cells were completely removed revealing the lattice network of connective tissue fibers within the alveolar surface. The method allows the complete removal of cellular elements of the lung while retaining the very fine 3D structure of the connective tissue matrix.  相似文献   

8.
L.K. Ives  A.W. Ruff 《Wear》1978,46(1):149-162
Scanning and transmission electron microscopy methods have been employed to study topographic features and subsurface damage associated with erosive-particle impact craters in annealed 310 stainless steel surfaces. Angular Al2O3 and spherical glass particles approximately 50 μm in diameter were projected at a velocity of 59 m s?1 to impact the surface at attack angles of 90° and 20°. Under these conditions, material was found to be displaced but not removed from the surface at isolated impact sites. A comparison was made with damage produced at diamond pyramid hardness indentations. Substantial differences were not observed. In general, a high dislocation density zone a few microns wide was found to surround both impact craters and hardness indentations. The width of this zone varied according to the size and shape of the crater and the direction of particle motion. Deformation twinning occurred at some impact sites. The plastic strain associated with impact craters in 310 stainless steel and copper was also determined by a method that is based on an analysis of selected-area electron channelling patterns.  相似文献   

9.
A technique for performing precision measurements of critical dimensions (CD) on integrated circuit (IC) features using the scanning electron microscope (SEM) is presented. The method is based on obtaining true in situ differential SEM video signals of IC features. The technique is characterized by its high sensitivity to feature edges, and relative slopes. Results are presented on the application of this technique to SEM measurements of IC patterns in silicon, photoresist, and aluminum. The method affords the capability of accurate pattern alignment during SEM imaging preceding linewidth measurements.  相似文献   

10.
Modern morphological investigation requires the use of a variety of technological approaches and the employment of rigorous morphometric analysis for an adequate evaluation of the structural and ultrastructural features of a tissue or organ. The introduction of the technique of freeze-cracking of tissue to expose new surfaces has made it possible to quantitate the normal surface characteristics of the glomerular capillaries of the mammalian kidney. This report describes the techniques used for the preparation and quantitative assessment of normal glomerular endothelial morphology. The techniques of in vivo and in vitro vascular perfusion of kidneys as a method of fixation and the freeze-cracking of tissue are outlined in detail. In addition, a morphometric analysis of the endothelial surface characteristics are described and values are reported for the control rat and human kidneys from transplant donors.  相似文献   

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13.
Numerical simulations of energy filtering effects on backscattered electron images of semiconductor multilayers are reported. The theoretical investigation has been performed for a wide range of energies, 1-40 keV, and for beam incidence angles between 90 degrees (normal incidence) and 20 degrees. Quite a general purpose of this research concerns the investigation of the optimum energy conditions and of their implications. It will be shown that the optimum energy defines an operating context suitable to ensure a compositional contrast enhancement; i.e. a minimum threshold current and a maximum resolution, without energy filtering, independent of the beam incidence angle. This optimum energy, depending on the specimen and its details, is, however, of the order of a few keV or less for specimen details having a size of the order of few nm. When the performance of the electron gun does not allow to work at low energy it is necessary to operate at an energy higher than the optimum one, the energy filtering can produce positive effects. Yet in those circumstances there is an optimum energy loss window suitable to minimise the threshold current. It spreads from 10-30%, depending on the primary energy and size of the compositional detail, for normal incidence, to a few per cent for high incidence angles and high energy. The simulation results for these last conditions are in agreement with the well-known experimental results obtained with the low-loss methods.  相似文献   

14.
A method of microtomography of layered microstructures during detection of backscattered electrons in a scanning electron microscope is described. This method is based on the formation of layer-by-layer images hidden under the surfaces of microstructures using reflected electrons filtered within a narrow energy window. An improved deflector-type spectrometer with toroidal electrostatic sector electrodes is applied for microtomography and spectroscopy. To improve the sharpness and accuracy of separation of individual buried heteroboundaries, the modulation principle of video-signal detection is implemented.  相似文献   

15.
A technique using pure gallium metal as a replication material is reported for biological surface scanning electron microscopy (SEM). The technique first directly enables aquatic organisms in water to be replicated due to gallium's low melting point and, second, reproduces surface structures and images of the two-dimensional (2D) distribution of substances transferred from the original surface to the gallium surface due to gallium's high surface tension. An aquatic protozoan in water was directly replicated to show its typical surface structures. The technique was then used to visualize human hair surface structures and 2D transferred substance distribution using X-ray microanalysis.  相似文献   

16.
A method for unmounting entire resin-embedded samples for SEM observation is described. This technique is particularly useful when correlation of TEM and SEM images is desired for material that is no longer available for conventional SEM preparative procedures. Sample embedded in a variety of epoxy-type resins were trimmed of excess resin and placed in a concentrated solution of sodium methoxide. After complete dissolution of the resin, the tissue was washed in a graded series of sodium methoxide in methanol--benzene, transferred to acetone, critical point dried, mounted on stubs, and coated with gold-palladium. Upon viewing in the SEM, the tissue sample showed remarkable preservation of detail at relatively high magnifications.  相似文献   

17.
Cells were cultivated on transparent conductive substrates, glass slides coated with indium oxide; individual cells were marked with a diamond indentor. Cell cultures were frozen (–15°C), thawed, and then stained with fluorescent dyes to determine cell damage. The marked cells were examined by phase contrast, fluorescence, and Nomarski DIC microscopy. After aldehyde and osmium tetroxide fixation, the cell preparations were sequentially treated with tannic acid, uranyl acetate, and lead citrate. The same marked cell could be sequentially studied by light microscopy (LM; in water immersion conditions), scanning electron microscopy (SEM; after dehydration and critical point drying), and transmission electron microscopy (TEM; after embedding of cell samples in epoxy resin and laser marking of the cell previously marked with a diamond indentor). The method used ensures good preservation of cell morphology, cell surface relief, and intracellular structures. The treatment used renders the cells conductive and permitted SEM of uncoated culture cells on conductive substrates.  相似文献   

18.
A unified treatment of the resolution of three closely related techniques is presented: emission electron microscopy (particularly photoelectron microscopy, PEM), low energy electron microscopy (LEEM), and mirror electron microscopy (MEM). The resolution calculation is based on the intensity distribution in the image plane for an object of finite size rather than for a point source. The calculations take into account the spherical and chromatic aberrations of the accelerating field and of the objective lens. Intensity distributions for a range of energies in the electron beam are obtained by adding the single-energy distributions weighted according to the energy distribution function. The diffraction error is taken into account separately. A working resolution is calculated that includes the practical requirement for a finite exposure time, and hence a finite non-zero current in the image. The expressions for the aberration coefficients are the same in PEM and LEEM. The calculated aberrations in MEM are somewhat smaller than for PEM and LEEM. The resolution of PEM is calculated to be about 50 A, assuming conventional UV excitation sources, which provide current densities at the specimen of 5 x 10(-5) A/cm2 and emission energies ranging up to 0.5 eV. A resolution of about 70 A has been demonstrated experimentally. The emission current density at the specimen is higher in LEEM and MEM because an electron gun is used in place of a UV source. For a current density of 5 x 10(-4) A/cm2 and the same electron optical parameters as for PEM, the resolution is calculated to be 27 A for LEEM and 21 A for MEM.  相似文献   

19.
The application of color cathodoluminescent scanning electron microscopy (CCL-SEM) for qualitative luminescence analysis of cholesterol, bilirubin, and protein in human gallstones was demonstrated. Images of these deposits (cholesterol, bilirubin, and protein) were formed in real colors (blue—cholesterol, red, orange—bilirubin, yellow, green—protein) in accordance with the cathodoluminescent spectrum for each control material. The other method described for transmission electron microscopy (TEM) of ultra-thin sections provides more detailed characterization of the ultrastructure of cholesterol-containing regions and their spatial interrelations with bilirubin-containing regions. Using CCL-SEM combined with TEM permits the receipt of more complete information about the chemical composition and ultrastructure of gallstones and may lead to more effective understanding of the pathogenesis of cholesterol cholelithiasis.  相似文献   

20.
Biotinylated deoxyadenosine triphosphate (dATP) (Bio-7-dATP) and 3H deoxythymidine triphosphate (dTTP) labeled adenovirus DNA were hybridized in situ to thin sections of Lowicryl K4M-embedded and whole-mount extracted HeLa cells infected with adenovirus. The biotinylated probe was detected by exposing the extracted cells or sections to antibodies against biotin followed by colloidal gold-conjugated secondary antibodies and then critical-point dried while 3H-dTTP labeled probe by electron microscopic autoradiography. On Lowicryl K4M sections, gold particles and silver grains were mainly restricted in the nucleus. Furthermore, whole-mount results suggested that replicating adenovirus DNA is localized on the nuclear matrix of its host cell. In this paper, the described non-radioactive procedures for hybrid detection offered several advantages: a) rapid signal detection; b) superior morphological preservation and spatial resolution; c) precise localization; and d) on Lowicryl K4M sections, signal to noise equivalent to radiolabeling.  相似文献   

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