scholarly journals Tilt-less 3-D electron imaging and reconstruction of complex curvilinear structures

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Emad Oveisi ◽  
Antoine Letouzey ◽  
Duncan T. L. Alexander ◽  
Quentin Jeangros ◽  
Robin Schäublin ◽  
...  
Author(s):  
Etienne de Harven

Biological ultrastructures have been extensively studied with the scanning electron microscope (SEM) for the past 12 years mainly because this instrument offers accurate and reproducible high resolution images of cell shapes, provided the cells are dried in ways which will spare them the damage which would be caused by air drying. This can be achieved by several techniques among which the critical point drying technique of T. Anderson has been, by far, the most reproducibly successful. Many biologists, however, have been interpreting SEM micrographs in terms of an exclusive secondary electron imaging (SEI) process in which the resolution is primarily limited by the spot size of the primary incident beam. in fact, this is not the case since it appears that high resolution, even on uncoated samples, is probably compromised by the emission of secondary electrons of much more complex origin.When an incident primary electron beam interacts with the surface of most biological samples, a large percentage of the electrons penetrate below the surface of the exposed cells.


Author(s):  
H.A. Cohen ◽  
W. Chiu ◽  
J. Hosoda

GP 32 (molecular weight 35000) is a T4 bacteriophage protein that destabilizes the DNA helix. The fragment GP32*I (77% of the total weight), which destabilizes helices better than does the parent molecule, crystallizes as platelets thin enough for electron diffraction and electron imaging. In this paper we discuss the structure of this protein as revealed in images reconstructed from stained and unstained crystals.Crystals were prepared as previously described. Crystals for electron microscopy were pelleted from the buffer suspension, washed in distilled water, and resuspended in 1% glucose. Two lambda droplets were placed on grids over freshly evaporated carbon, allowed to sit for five minutes, and then were drained. Stained crystals were prepared the same way, except that prior to draining the droplet, two lambda of aqueous 1% uranyl acetate solution were applied for 20 seconds. Micrographs were produced using less than 2 e/Å2 for unstained crystals or less than 8 e/Å2 for stained crystals.


Author(s):  
Kazumichi Ogura ◽  
Michael M. Kersker

Backscattered electron (BE) images of GaAs/AlGaAs super lattice structures were observed with an ultra high resolution (UHR) SEM JSM-890 with an ultra high sensitivity BE detector. Three different types of super lattice structures of GaAs/AlGaAs were examined. Each GaAs/AlGaAs wafer was cleaved by a razor after it was heated for approximately 1 minute and its crosssectional plane was observed.First, a multi-layer structure of GaAs (100nm)/AlGaAs (lOOnm) where A1 content was successively changed from 0.4 to 0.03 was observed. Figures 1 (a) and (b) are BE images taken at an accelerating voltage of 15kV with an electron beam current of 20pA. Figure 1 (c) is a sketch of this multi-layer structure corresponding to the BE images. The various layers are clearly observed. The differences in A1 content between A1 0.35 Ga 0.65 As, A1 0.4 Ga 0.6 As, and A1 0.31 Ga 0.69 As were clearly observed in the contrast of the BE image.


Author(s):  
D. R. Liu ◽  
S. S. Shinozaki ◽  
J. S. Park ◽  
B. N. Juterbock

The electric and thermal properties of the resistor material in an automotive spark plug should be stable during its service lifetime. Containing many elements and many phases, this material has a very complex microstructure. Elemental mapping with an electron microprobe can reveal the distribution of all relevant elements throughout the sample. In this work, it is demonstrated that the charge-up effect, which would distort an electron image and, therefore, is normally to be avoided in an electron imaging work, could be used to advantage to reveal conductive and resistive zones in a sample. Its combination with elemental mapping can provide valuable insight into the underlying conductivity mechanism of the resistor.This work was performed in a CAMECA SX-50 microprobe. The spark plug used in the present report was a commercial product taken from the shelf. It was sectioned to expose the cross section of the resistor. The resistor was known not to contain the precious metal Au as checked on the carbon coated sample. The sample was then stripped of carbon coating and re-coated with Au.


Author(s):  
J.R. Pfeiffer ◽  
J.C. Seagrave ◽  
C. Wofsy ◽  
J.M. Oliver

In RBL-2H3 rat leukemic mast cells, crosslinking IgE-receptor complexes with anti-IgE antibody leads to degranulation. Receptor crosslinking also stimulates the redistribution of receptors on the cell surface, a process that can be observed by labeling the anti-IgE with 15 nm protein A-gold particles as described in Stump et al. (1989), followed by back-scattered electron imaging (BEI) in the scanning electron microscope. We report that anti-IgE binding stimulates the redistribution of IgE-receptor complexes at 37“C from a dispersed topography (singlets and doublets; S/D) to distributions dominated sequentially by short chains, small clusters and large aggregates of crosslinked receptors. These patterns can be observed (Figure 1), quantified (Figure 2) and analyzed statistically. Cells incubated with 1 μg/ml anti-IgE, a concentration that stimulates maximum net secretion, redistribute receptors as far as chains and small clusters during a 15 min incubation period. At 3 and 10 μg/ml anti-IgE, net secretion is reduced and the majority of receptors redistribute rapidly into clusters and large aggregates.


Author(s):  
R.F. Stump ◽  
J.R. Pfeiffer ◽  
JC. Seagrave ◽  
D. Huskisson ◽  
J.M. Oliver

In RBL-2H3 rat basophilic leukemia cells, antigen binding to cell surface IgE-receptor complexes stimulates the release of inflammatory mediators and initiates a series of membrane and cytoskeletal events including a transformation of the cell surface from a microvillous to a lamellar topography. It is likely that dynamic properties of the IgE receptor contribute to the activation of these responses. Fewtrell and Metzger have established that limited crosslinking of IgE-receptor complexes is essential to trigger secretion. In addition, Baird and colleagues have reported that antigen binding causes a rapid immobilization of IgE-receptor complexes, and we have demonstrated an apparent increase with time in the affinity of IgE-receptor complexes for antigen.


Author(s):  
Allen Angel ◽  
Kathryn A. Jakes

Fabrics recovered from archaeological sites often are so badly degraded that fiber identification based on physical morphology is difficult. Although diagenetic changes may be viewed as destructive to factors necessary for the discernment of fiber information, changes occurring during any stage of a fiber's lifetime leave a record within the fiber's chemical and physical structure. These alterations may offer valuable clues to understanding the conditions of the fiber's growth, fiber preparation and fabric processing technology and conditions of burial or long term storage (1).Energy dispersive spectrometry has been reported to be suitable for determination of mordant treatment on historic fibers (2,3) and has been used to characterize metal wrapping of combination yarns (4,5). In this study, a technique is developed which provides fractured cross sections of fibers for x-ray analysis and elemental mapping. In addition, backscattered electron imaging (BSI) and energy dispersive x-ray microanalysis (EDS) are utilized to correlate elements to their distribution in fibers.


Author(s):  
Paul J. Wright

Most industrial and academic geologists are familiar with the beautiful red and orange cathodoluminescence colours produced by carbonate minerals in an optical microscope with a cold cathode electron gun attached. The cement stratigraphies interpreted from colour photographs have been widely used to determine the post depositional processes which have modified sedimentary rock textures.However to study quartzose materials high electron densities and kV's are necessary to stimulate sufficient emission. A scanning electron microscope with an optical collection system and monochromator provides an adequate tool and gives the advantage of providing secondary and backscattered electron imaging as well as elemental analysis and distribution mapping via standard EDS/WDS facilities.It has been known that the incorporation of many elements modify the characteristics of the CL emissions from geological materials. They do this by taking up positions between the valence and conduction band thus providing sites to assist in the recombination of electron hole pairs.


Author(s):  
D. R. Liu ◽  
D. B. Williams

The secondary electron imaging technique in a scanning electron microscope (SEM) has been used first by Millman et al. in 1987 to distinguish between the superconducting phase and the non-superconducting phase of the YBa2Cu3O7-x superconductors. They observed that, if the sample was cooled down below the transition temperature Tc and imaged with secondary electrons, some regions in the image would show dark contrast whereas others show bright contrast. In general, the contrast variation of a SEM image is the variation of the secondary electron yield over a specimen, which in turn results from the change of topography and conductivity over the specimen. Nevertheless, Millman et al. were able to demonstrate with their experimental results that the dominant contrast mechanism should be the conductivity variation and that the regions of dark contrast were the superconducting phase whereas the regions of bright contrast were the non-superconducting phase, because the latter was a poor conductor and consequently, the charge building-up resulted in high secondary electron emission. This observation has since aroused much interest amoung the people in electron microscopy and high Tc superconductivity. The present paper is the preliminary report of our attempt to carry out the secondary electron imaging of this material in a scanning transmission electron microscope (STEM) rather than in a SEM. The advantage of performing secondary electron imaging in a TEM is obvious that, in a TEM, the spatial resolution is higher and many more complementary techniques, e.g, diffraction contrast imaging, phase contrast imaging, electron diffraction and various microanalysis techniques, are available.


Author(s):  
Etienne de Harven ◽  
Hilary Christensen ◽  
Richard Leung ◽  
Cameron Ackerley

The T-derived subset of human peripheral blood normal lymphocytes has been selected as a model system to study the usefulness of 5 nm gold markers for quantification of single epitopes expressed on cell surfaces. The chosen epitopes are parts of the CD3 and CD5 molecules and can be specifically identified by hybridoma produced monoclonal antibodies (MoAbs; LEU-4 and LEU-1; Becton-Dick- inson, Mountain view, CA) . An indirect immunolabeling procedure, with goat anti-murine IgG adsorbed on the surface of 5 nm colloidal gold particles (GAM-G5, Janssen Pharmaceutica, Beerse, Belgium) has been used. Backscattered Electron Imaging (BEI) in a field emission scanning electronmicroscope (SEM) and transmission electron microscopy of thin sections of lymphocytes labeled before plastic embedding, were both used to identify and quantitate gold labeled cell surface sites, Estimating that the thickness of “silver” sections is approximately 60 nm and counting the number of gold particles on the entire cell perimeter, we calculated that, for LEU-4, the number of markers per um2 of cell surface is in the 140-160 range (Fig.l). Cell contour length measurements indicated that the surface of one lymphocyte is approximately 130-160 um2 that of a smooth sphere of identical diameter, reflecting the role of microvilli in expanding the surface area. The total number of gold labeled sites on the surface of one lymphocyte averages, therefore between 20,000 and 24,000 per cell.


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