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  • Micro-CT of Insect Larva Protocol

    Species: red blood worm (midge larva) Critical point drying of midge larvae with subsequent X-ray micro-computed tomography (micro-CT) to reconstruct the inner anatomy.
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  • Contrast Enhancement of Polycrystalline Metals - Sample Preparation for SEM

    Application Note - Ion milling is a perfect alternative for chemical etching, especially for polycrystalline metals, such as copper. Ion milling can be used to increase the contrast of the grain structure and their interfaces. In contrast to chemical etching the milling process is clean, safe and easy to operate. Ion energy and milling time depend on the milling rate of the metal.
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  • Drosophila larvae - Sample Preparation for Cryo-SEM

    Application Note for Leica EM ACE900 - Drosophila larvae were sandwiched between two 3 mm aluminum slit carriers with the 100 μm cavities facing each other and high-pressure frozen with a Leica EM HPM100. No ethanol as synchronization media was used, 1-hexadecene was used as filler. The wholes of the slit carriers were filled with filter tips dipped in 1-hexadecene to keep the carrier sandwich complete after freezing.
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  • Giardia lamblia - Sample Preparation for Cryo-SEM

    Application Note for Leica EM ACE900 - A 100 mesh copper grid (12 um thickness) was dipped into a concentrated Giardia suspension and sandwiched between two flat 3 mm aluminum specimen carriers with scratched surfaces. Subsequently, the sandwich was transferred to the widened hole of a middle plate (3.1 mm diameter). A 50 um spacer ring was added on top and the specimen immediately frozen with an HPM100 high-pressure freezing machine without using alcohol as synchronization fluid.
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  • Cross Sectioning of Cadmiumsulphide (CdS) for Cathodoluminescence

    Cathodoluminescence can be used to achieve spectra and high resolution images of impurity and structural defects in semicondoctors, minerals and insulating materials. This application note explains how to prepare a perfect sample surface for carhodoluminescence and how to use ion beam slope cutting to prepare the sample surface free of any preparation artefacts.
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  • Picea abies (L.) KARST - Sample Preparation for TEM

    Application Note for Leica EM AMW - Plants (5-years old) were grown in pots filled with soil and kept in greenhouse conditions. Five weeks before harvesting the plants were transferred into growth chambers and cultivated at a temperature of 20°C during daytime and 12°C overnight. The relative humidity was set at 60% and the photoactive radiation was 500 μmol m-2 s-1 during daytime. Sample preparation for transmission electron microscopy (TEM) was performed in order to develop a standard protocol that would reduce sample preparation time for TEM-investigations. Therefore the overall and fine structure of leaf cells prepared with the Leica EM AMW were compared with leaf cells that were prepared with a conventional fixation protocol at room temperature.
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  • Cross Sectioning of Copper for Electron Backscattered Diffraction (EBSD)

    Application Note for Leica EM TIC 3X - Electron Backscattered Diffraction (EBSD) is a surface technique creating diffraction patterns (Kikuchi-bands). It can be used for crystal orientation mapping, defect studies, phase identification, grain boundary studies and morphological studies. The information depth is just a few nm, therefore good sample preparation is very important to avoid damages.
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  • Porous Ceramics - Sample Preparation for SEM

    Application Note for Leica EM RES102 - Ceramic membrane filters with pore sizes down to a few nanometres must be investigated in cross-section with regard to the structure of the pores. The smallest pores are of special interest. In most cases, conventional grinding methods cannot be used for such problems, as the pore structure would be distorted. This applies in particular to the pores in the nanometre range.
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  • Maple (Acer saccharum) Leaves - High Pressure Freezing and Freeze Substitution for TEM

    Application Note for Leica EM HPM100 - Leaves were immersed in hexadecene and placed under a gentle (0.3 bar) vacuum for 10 minutes to evacuate the internal air spaces. The leaves were then trimmed to fit the carriers and placed in the 200 μm side of a 6 mm Type A specimen carrier. Free space was filled with additional hexadecene after which a 6 mm Type B specimen carrier was placed on top with the flat side down.
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  • Removal of Surface Layers - Sample Preparation for SEM and TEM

    Application Note for Leica EM RES102 - Sometimes it is necessary to remove surface layers to gain access to the real surface structure. That can be a native oxide, or layers coming from the preparation process itself, like re-deposition. Depending on the layers thickness and the energy used for the cleaning process, it takes between a few seconds and half an hour. The energy depends on the milling rate of the material.
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  • Semiconductor Structures with Large Differences in Hardness - Sample Preparation for SEM

    Application Note for Leica EM RES102 - In most cases, multi-layer structures or material combinations with large differences in hardness cannot be processed with conventional polishing techniques, or can only be very poorly processed. Due to the large differences in hardness, blurring or edge-rounding occurs, which distorts the original structure. In the application example shown, we are dealing with a solder ball structure. The goal was to determine the solder structure. The use of conventional grinding and polishing techniques was not possible due to the large differences in hardness between the solder ball and other materials in the sample.
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  • Cross Section of Solar Cells

    Application Note for Leica EM TIC020, Leica EM TIC 3X - Cross section of a complete solar cell.
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  • Cross Sectioning of a Multilayer System - Preparation of a Perfect Sample Surface for EBSD

    Application Note for Leica EM TIC 3X - Electron Backscattered Diffraction (EBSD) is a surface technique creating a diffraction pattern (Kikuchi-bands). It can be used for crystal orientation mapping, defect studies, phase identification, grain boundary studies and morphology studies. The information depth is just a few nm. Therefore good sample preparation is very important to avoid any damage. This is very difficult in case of multilayer system with big differences in hardness.
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  • High-Pressure Freezing and Freeze Substitution of Hep-2 Cells Infected with Chlamydia pneumoniae

    Application Note for Leica EM HPM100 - Hep-2 cells infected with Chlamydia pneumoniae were cultured on carbon-coated 6 mm Sapphire discs. Cells were high-pressure frozen in an EM HPM100 using the 6 mm CLEM middle plate with following setup: Sapphire disc with cells, spacer 200 μm, bare Sapphire disc, 2 spacers 200 μm. Ethanol was used as a synchronization fluid to transfer pressure at room temperature prior to cooling.
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  • Multilayer Systems with Widely Different Sputter Rates - Sample Preparation for TEM

    Application Note for Leica EM RES102 - The multi-layer system to be prepared in cross-section consists of a Si substrate, a TiN layer with a thickness of a few nm and a 500 nm W layer. All these components have extreme differences in their hardness, their atomic weight and in their sputter rates. A preparation of this kind of samples with sample rotation would lead to a wall overlying the area of the layers.
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  • In-Containing Compound Semiconductors - Sample Preparation for TEM

    Application Note for Leica EM RES102 - Previous studies showed that surface accumulation of In occurs when InP was milled in a conventional way with Ar ions. The consequence is In islands on the sample surface. This leads to low quality of TEM samples. To remove these islands, reactive ion milling with iodine ions (RIBE / CAIBE) can be used. This method has the disadvantage of polluting the ion guns and the vacuum system of the ion milling device and leads to chemical reactions with the sample material. To avoid these problems we prepared these samples very gently with low energy Ar ions.
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  • Carbon Coating for Polymeric Materials

    Application Note fo Leica EM ACE600 - A solid understanding of polymer property-structure relationships is critical to improve and shorten development routes to new products. A direct way to determine correlations between structure and mechanical properties is provided by electron microscopy. Electron microscopy techniques have an important advantage over other methods, as they can provide local information at high spatial resolution. However, a major problem with polymers is their inherent lack of contrast.
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  • Commercially Available Hand Creams - Sample Preparation for Cryo-SEM

    Application Note for Leica EM HPM100 - Hand creams having different water contents were applied into the 100 μm cavities of two 3 mm type A sample carriers which were then closed cream sides inwards. The sample assembly was high pressure frozen with a Leica EM HPM100 and moved to a cooled Leica EM VCT100 loading station.
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  • "Shallow Trench Isolation" Structures - Sample Preparation for TEM

    Application Note for Leica EM RES102 - The cross-sectional preparation of structured semiconductor materials requires a very thorough mechanical pre-preparation. In doing this, it must be ensured that the structure of interest should be located as close to the centre of the sample as possible. As the sample will be ion milled from both sides, a specific preparation of the structure is necessary in most cases, which means that you must thin these structures from both sides.
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  • Tobacco Leaf - Critical Point Drying Protocol for SEM

    Application Note for Leica EM CPD300 - Critical point drying of tobacco leafs with subsequent platinum coating and SEM analysis.
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  • Paper Samples - Sample Preparation for SEM

    Application Note for Leica EM RES102 - A coated paper sample has been prepared with ion beam slope cutting in order to test the procedure with regard to its applicability. With the use of ion beam slope cutting a cross section of paper could be prepared. On the basis of this sample processing, it was possible to show the largely unaffected original structure of the thermally-sensitive paper in the scanning electron microscope.
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  • Metal Films and Sheets - Sample Preparation for TEM

    Application Note for Leica EM RES102 - Most metal films already have a thickness that requires no further mechanical pre-preparation. Frequently, however, they are also domed, which can lead to undefined milling angles. This is a disadvantage, particularly for films that contain inclusions, and that therefore mostly require very flat milling angles. Metal sheets are thicker than 100 µm. Mechanical pre-preparation is necessary to obtain an acceptable initial thickness and a good surface quality for ion milling.
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  • Nematode E. dianae - Critical Point Drying Protocol for SEM

    Application Note for Leica EM CPD300 - Critical point drying of nematode Eubostrichus dianae to detect the ectosymbiotic bacteria layer with subsequent gold coating ans SEM analysis.
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  • Visualization of Membrane Dynamics with Millisecond Temporal Resolution

    Application Note for Leica EM ICE, Leica EM AFS2 - Electrical stimulation of neurons combined with high-pressure freezing allows physiological activation of synaptic activity and precise control over the time frame of the induced synaptic activity.
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  • Cross-Sectional Preparation of Structured Semiconductor Materials for TEM

    Application Note for Leica EM RES102 - The vertical layer construction of a semiconductor structure should be examined as a TEM cross-sectional sample. In addition to the specific preparation of the desired structure, the widely different sputter rates and atomic weights of the individual components represent the level of difficulty involved with this preparation problem.
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  • Contrast Enhancement of Polished Cross Sections of Semiconductor Structures - Sample Preparation for SEM

    Application Note for Leica EM RES102 - The surfaces of polished cross sections often show fine scratches and residues of the removed material or of the abrasive material. The artefacts are strongly material-dependent, and are mostly only detectable at higher resolutions in the scanning electron microscope. A further problem arises from the fact that the ground section mostly only has low contrast, i.e., in the structures of the semiconductor materials are very difficult to discern. With the use of ion beam milling, the ground sections of semiconductor structures can be "contrasted".
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  • Wall Cress Pod Protocol - Critical Point Drying of Arabidopsis thaliana for SEM

    Application Note for Leica EM CPD300 - Critical point drying of wall cress (Arabidopsis thaliana) pod with subsequent gold coating and SEM analysis.
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  • Surface Modification of ZnAg Sample - Sample Preparation for SEM

    Application Note for Leica EM RES102 - By means of cleaning, polishing and contrast enhancement a soft ZnAg sample should be prepared to obtain information concerning the grain structure and interfaces of the sample. The sample is contaminated after mechanical polishing. There are still some scratches on the surface. Grain structure is almost invisible.
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  • Cross Sectioning of Ni/Cu on Steel for EBSD

    Application Note for Leica EM TIC 3X - Electron Backscattered Diffraction (EBSD) is a surface technique creating a diffraction pattern (Kikuchi-bands). It can be used for crystal orientation mapping, defect studies, phase identification, grain boundary studies and morphology studies. The information depth is just a few nm. Therefore a good sample preparation is very important.
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  • Bacteria Protocol - Critical Point Drying of E. coli for SEM

    Application Note for Leica EM CPD300 - Critical point drying of E. coli with subsequent platinum / palladium coating and SEM analysis. Sample was inserted into a filter disc (Pore size: 16 - 40 μm) and placed into the filter discs and porous pots holder. Cultivate fungi and bacteria on agar containing growth medium for 3 days.
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