Specimen Preparation For Electron Microscopy Research Articles

The preparation of transmission electron microscopy specimens of as-drawn gold wire

The preparation of transmission electron microscopy specimens of as-drawn gold wire

New Technique for Successful Thermal Barrier Coating Specimen Preparation for Transmission Electron Microscopy

Reliability of thermal barrier coatings (TBC) hinges on the adhesion of a thermally grown oxide scale to an insulative ceramic topcoat and an underlying metallic bondcoat. The width of the scale and its interfaces makes transmission electron microscopy (TEM) an appropriate tool for its analysis. However, specimen preparation has proven to be a challenging obstacle leading to a dearth of TEM research on TBCs. A new approach to cross-section TBC TEM specimen preparation is described. The principal advantages of this technique are reproducibility, reduced specimen damage, and time savings resulting from decreased ion milling. This technique has been successfully applied to numerous TBC specimens with various thermal histories.

Biological Specimen Preparation for Transmission Electron Microscopy. Audrey M. Glauert , Peter R. Lewis

Previous articleNext article No AccessNew Biological BooksBiological Specimen Preparation for Transmission Electron Microscopy. Audrey M. Glauert , Peter R. Lewis Sara E. MillerSara E. Miller Search for more articles by this author PDFPDF PLUS Add to favoritesDownload CitationTrack CitationsPermissionsReprints Share onFacebookTwitterLinkedInRedditEmail SectionsMoreDetailsFiguresReferencesCited by The Quarterly Review of Biology Volume 75, Number 1Mar., 2000 Published in association with Stony Brook University Article DOIhttps://doi.org/10.1086/393374 Views: 1Total views on this site Copyright 2000 The University of ChicagoPDF download Crossref reports no articles citing this article.

Unexpected property of trehalose as observed by cryo-electron microscopy.

Trehalose is an agent useful in maintaining the integrity of many biological systems submitted to various stresses. It is also presumed to improve specimen preparation for electron microscopy and to reduce beam damage. Here we study the effect of trehalose on the preparation and observation by cryo-electron microscopy of thin vitrified films of biological suspensions. We observe that trehalose, as compared to sucrose, can indeed reduce electron beam damage to biological particles, as determined from the dose necessary for the onset of bubbling. Surprisingly, we also find that the contrast of biological particles is higher in a vitrified solution of trehalose than in one of sucrose. This effect can be explained if the water evaporation during the specimen preparation is less in the presence of trehalose than with sucrose, but we do not yet understand the underlying reasons since the evaporation properties of both sugars are similar at a macroscopic level. We conclude that trehalose is truly a remarkable substance and that more investigation is needed in order to fully understand its properties, and that the addition of ca. 3-5% trehalose to biological suspensions is a simple and useful method to reduce commonly arising drying artefacts and water evaporation in the thin film vitrification method.

Biological specimen preparation for transmission electron microscopy

Biological specimen preparation for transmission electron microscopy

Method for Cross-sectional Transmission Electron Microscopy Specimen Preparation of Composite Materials Using a Dedicated Focused Ion Beam System.

: A new method for transmission electron microscope (TEM) specimen preparation using a focused ion beam (FIB) system that results in a lower rate of gallium (Ga) implantation has been developed. The method was applied to structural and analytical studies of composite materials such as silicon (Si)-devices and magneto-optical disk. To protect the specimens against Ga ion irradiation, amorphous tungsten (W) was deposited on the surface of the specimen prior to FIB milling. The deposition was quite effective in reducing the Ga implantation rate, and energy-dispersive X-ray (EDX) analysis of these specimens detected 0.3-1.5% Ga incorporated in the thinned area. FIB milling times for these specimens were 1.5-2 hr, with a starting thickness of about 50 µm. Although the milling rate was high, all the materials were properly prepared for TEM study, and clear crystal lattice images were observed on all specimens.

TEM FIB Lift-Out of Mount Saint Helens Volcanic Ash

Focused Ion Beam (FIB) specimen preparation for both scanning and transmission electron microscopy (SEM and TEM respectively) has seen an increase in usage over the past few years. The advantage to the FIB is that site specific cross sections (or plan view sections) may be fabricated quickly and reproducibly from numerous types of materials using a finely focused beam of Ga+ ions [1,2]. It was demonstrated by Prenitzer et al. that TEM specimens may be acquired from individual Zn powder particles by employing the FIB LO specimen preparation technique [3]. In this paper, we use the FIB LO technique to prepare TEM specimens from Mount Saint Helens volcanic ash.Volcanic ash from Mount Saint Helens was obtained at the Microscopy and Microanalysis 1998 meeting in Atlanta. TEM analysis of the ash was performed using the FIB lift out technique [1]. Ash powders were dusted onto an SEM sample stud that had been coated with silver paint.

A Tutorial of the Fib Lift-Out Technique for TEM Specimen Preparation

Abstract The focused ion beam (FIB) instrument has been developed and exploited by the microelectronics arena for specimen preparation for both scanning and transmission electron microscopy (TEM). The inception [1] and subsequent development [2] of the FIB TEM lift-out (LO) technique has enabled electron transparent membranes of generally uniform thickness to be produced for TEM analysis. The primary advantage of the FIB technique is that site specific cross sections (or plan view sections [3]) may be fabricated quickly and reproducibly. The FIB LO technique has been used extensively in our laboratory for a wide range of materials [4] and biological applications [5] which are summarized in figure 1. The FIB LO method consists of milling a series of trenches around an area of interest. Then the bulk sample is tilted up to ∼60 degrees to allow the beam to impinge on the lower portion of the specimen surface so that cuts can be made along the bottom edge and the lower 2/3 of the distance up one side of the specimen.

A review of focused ion beam milling techniques for TEM specimen preparation

The use of focused ion beam (FIB) milling for the preparation of transmission electron microscopy (TEM) specimens is described. The operation of the FIB instrument is discussed and the conventional and lift-out techniques for TEM specimen preparation and the advantages and disadvantages of each technique are detailed. The FIB instrument may be used for rapid site-specific preparation of both cross-section and plan view TEM specimens.

Wetting of copper on α-Al 2O 3 surfaces depending on the orientation and oxygen partial pressure

The influence of oxygen partial pressure and misorientation on the chemistry and bonding at interfaces between Cu and single crystalline α-Al 2O 3 was studied by analytical electron microscopy with nanoscale resolution. The interfaces were prepared by the sessile drop technique, i.e. by melting pure Cu (99.99%) in a furnace onto α-Al 2O 3 plates. We investigated the chemistry and bonding at the interfaces for two orientations of the single crystalline α-Al 2O 3 substrates with two different surface planes, the basal (B-) plane, (0001), α-Al 2O 3 B, and the rhombohedral (R-) plane, (101̄2), α-Al 2O 3 R. The oxygen partial pressure ( p Ox) in the furnace was varied between 10 −4 and 10 −15 bar. This was controlled using a solid state electrolyte cell instrument. Spatially resolved (SR) electron energy-loss spectroscopy (EELS) in a dedicated scanning transmission electron microscope (STEM) was applied to characterize the chemistry and the electronic structure at the interfaces. At low oxygen partial pressure ( p Ox<3×10 −11 bar), all interfaces fractured during transmission electron microscopy (TEM) specimen preparation, independently of the α-Al 2O 3 surfaces (B- or R-plane). At higher oxygen partial pressures (p Ox =[1.5×10 −10…7.8×10 −4] bar) the successful TEM specimen preparation indicates stronger bonding across the interfaces. No changes in the electronic structure at the Cu/ α-Al 2O 3 B interfaces as compared to the bulk were found by spatially resolved EELS while the electronic structure at the Cu/ α-Al 2O 3 R interfaces changed strongly. These differences are attributed to the reconstruction of the α-Al 2O 3 B plane being neutral compared to the relaxation of the α-Al 2O 3 R-plane which is charged and can therefore form ionic bonds with the Cu.

High-pressure freezing for preservation of high resolution fine structure and antigenicity for immunolabeling.

What is high-pressure freezing (HPF)? HPF is a method of specimen preparation for electron microscopy (EM) that freezes noncryoprotected samples up to 0.5 mm in thickness without significant ice crystal damage. Other freezing methods are limited to 1/100 (plunge and impact freezing) to 1/10 (propane jet freezing) that sample thickness. With HPF you are working in the realm of whole cells and tissues instead of small cells or cell cortical regions. For more details on the theory of how HPF works, and some examples of its application, there are several reviews available (-).

Transmission electron microscope specimen preparation of Zn powders using the focused ion beam lift-out technique

Particles of Zn powder have been studied to show that high-quality scanning electron microscope (SEM) and transmission electron microscope (TEM) specimens can be rapidly produced from a site-specific region on a chosen particle by the focused ion beam (FIB) lift-out technique. A TEM specimen approximately 20-µm long by 5-µm wide was milled to electron transparency, extracted from the bulk particle, and micromanipulated onto a carbon coated copper mesh TEM grid. Using the FIB lift-out method, we were able to prepare a site-specific TEM specimen from a difficult material in under 3 hours. The TEM analysis of the lift-out specimen revealed a large amount of thin area free from characteristic signs of damage that may be observed as a result of conventional argon ion milling. The overall microstructure of the specimen prepared by the FIB lift-out method was consistent with samples prepared by conventional metallographic methods. A grain size of ∼10 to 20 µm was observed in all specimens by both TEM and SEM analysis. Light optical microscopy revealed the presence of internal voids in ∼10 to 20 pct of all particles. The SEM analysis showed the voids to extend over ∼70 pct of the particle volume in some cases.

Substrate, Inhibitor, or Antibody Stabilizes the Glu 119 Gly Mutant Influenza Virus Neuraminidase

We have previously reported the isolation and characterization of an influenza virus variant with decreased sensitivity to the neuraminidase-specific inhibitor zanamivir. This variant, which has a mutation in the active site, Glu 119 Gly (E119G), has the same specific activity as the wild-type neuraminidase (NA), but is inherently unstable, as measured by loss of both enzyme activity and NC10 monoclonal antibody reactivity. However, despite the instability of the NA, replication of the virus in liquid culture is not adversely affected. We demonstrate here that in addition to enhanced temperature sensitivity the mutant NA was significantly more sensitive to formaldehyde and to specimen preparation for electron microscopy. Substrate, inhibitor, or monoclonal antibodies stabilized the NA against all methods of denaturation. These results suggest that the instability of the variant is primarily at the level of polypeptide chain folding rather than at the level of association of monomers into tetramers. Furthermore the presence of high levels of substrate, either cell or virus associated, may be sufficient to stabilize the NA during virus replication.

Applications of the FIB lift-out technique for TEM specimen preparation.

A site-specific technique for cross-section transmission electron microscopy specimen preparation of difficult materials is presented. A focused ion beam was used to slice an electron transparent membrane from a specific area of interest within a bulk sample. Micromanipulation lift-out procedures were then used to transport the electron-transparent specimen to a carbon-coated copper grid for subsequent TEM analysis. The FIB (focused ion beam) lift-out technique is a fast method for the preparation of site-specific TEM specimens. The versatility of this technique is demonstrated by presenting cross-sectioned TEM specimens from several types of materials systems, including a multi-layered integrated circuit on a Si substrate, a galvanized steel, a polycrystalline SiC ceramic fiber, and a ZnSe optical ceramic. These specimens have both complex surface geometry and interfaces with complex chemistry. FIB milling was performed sequentially through different layers of cross-sectioned materials so that preferential sputtering was not a factor in preparing TEM specimens. The FIB lift-out method for TEM analysis is a useful technique for the study of complex materials systems for TEM analysis.

A plasma-polymerized protective film for transmission electron microscopy specimen preparation by focused ion beam etching

Focused ion beam (FIB) milling has been widely used for the preparation of cross-sectional transmission electron microscopy (X-TEM) specimens. In FIB fabrication, a protective layer is normally formed on the specimen surface by ion-beam-assisted metal deposition to avoid the damage caused by highly accelerated ion beams. However, the metal deposition itself induces damage in the specimen. This problem is especially serious when the region near the surface of a specimen is of interest. Investigation of subsurface damage through X-TEM observations showed that, for 30 keV FIB, the defects induced by ion-beam-assisted deposition extend about 400 Å from the surface. In the case of single-crystalline silicon, a several-hundred-angstrom layer just below the surface is converted into an amorphous layer. We propose the use of a plasma-polymerized organic film which is formed by plasma polymerizing of methane and ethylene, instead of the metal deposit. We used the film as a protective layer and fabricated TEM specimens without damaging the fine subsurface structures. In the case of single-crystalline silicon, no amorphous layer was observed. In addition, a subsurface structure in an aluminum sample prepared by the FIB method was successfully observed by TEM.

Protocol of Plant Specimen Preparation for Transmission Electron Microscopy (Conventional Methods)

Abstract These notes address conventional specimen preparation for higher plant tissues. While in many cases freeze-substitution has been shown to be a superior method, the equipment required is not generally available in most electron microscopic laboratories, Thus, the more conventional procedures outlined herein have been found to work satisfactorily for most leaf material, and have been most successful in the fixation and embedment of small root slices (from near the root tip), An adjustment of the osmotic pressure may be necessary if the plant material used is highly vacuolated and if there is an exceptionally high water content.

Focused Ion Beam Milling for Site Specific Scanning and Transmission Electron Microscopy Specimen Preparation

It has been shown that a focused ion beam (FIB) instrument may be used to prepare site specific cross-sectioned specimens to within &lt; 0.1 μm for both scanning and transmission electron microscopy (SEM and TEM, respectively). FIB specimen preparation has been used almost exclusively in the microelectronics industry. Recently, FIB specimen preparation has been utilized for other materials systems and applications.A cross-sectioned SEM specimen is produced by sputtering away a trench of material from near the region of interest. Large amounts of material are sputtered using large ion beam diameters (e.g., l00’s nm) and high beam current (e.g., l000’s pA), while the final sputtering operations are achieved using smaller beam diameters (e.g., &lt; 10 nm) and lower beam current (e.g., 10’s of pA). The SEM specimen may then be etched to reveal particular microstructural features of interest. A low magnification SEM image of a multi-layered device prepared for cross-section analysis by the FIB method is shown in FIG. 1.

A method to study the effect of chemical dissolution on the morphology of soil clay

Soil clays contain a relatively large amount of disordered inorganic material. Chemical dissolution has been used for the removal of this material (Jackson, 1956; Hashimoto &amp; Jackson, 1960; Follett et al, 1965 a,b; Wada &amp; Greenland, 1970). On the other hand, Farmer et al. (1977) claimed that dissolution of allophane and imogolite with hot 5% Na2CO3 for periods of 2-100 h led to new phases which could be distinguished from the starting material by infrared spectroscopy. It is clear, therefore, that chemical dissolution can alter soil clays. This note suggests an electron microscopy specimen preparation technique to study the morphological changes. Collodion films containing densely arranged minute hollows are used for specimen supports.

Transmission Electron Microscopy Specimen Preparation of Ceramic Coatings on Ceramic Fibers

AbstractA method of preparing transmission electron microscopy (TEM) specimens of coated ceramic fibers has been developed that produces large areas containing many electron transparent fibers, due to the minimal preferential milling of the epoxy matrix. Multiple individual fibers or tows are impregnated with a high-temperature epoxy and contained to assure a high fiber-to-epoxy volume ratio. The samples are then sectioned and mechanically thinned either parallel or normal to the fiber axes using a wedge polisher on diamond lapping films to achieve a thickness of less than 5 μm. The thinned sample is then ion-milled to electron transparency in less than 30 min, giving representative specimens of the coating, fiber, and coating-fiber interface. This technique is also well suited to preparing extremely flat specimens for both light microscopy and scanning electron microscopy analysis of thin coatings.

Focused Ion Beam Milling and Micromanipulation Lift-Out for Site Specific Cross-Section Tem Specimen Preparation

AbstractA site specific technique for cross-section transmission electron microscopy specimen preparation of difficult materials is presented. Focused ion beams are used to slice an electron transparent sliver of the specimen from a specific area of interest. Micromanipulation lift-out procedures are then used to transport the electron transparent specimen to a carbon coated copper grid for subsequent TEM analysis. The experimental procedures are described in detail and an example of the lift-out technique is presented.