Leica Science Lab - Tag : Sample Preparation https://www.leica-microsystems.com//science-lab/tag/sample-preparation/ Article tagged with Sample Preparation en-US https://www.leica-microsystems.com/71898 EM Sample Preparation Fast, high-quality vitrification with the EM ICE High Pressure Freezer The EM ICE High Pressure Freezer was developed with a unique freezing principle and uses only a single pressurization and cooling liquid: liquified nitrogen (LN2). This design enables three major benefits that lead to high-quality sample vitrification results. https://www.leica-microsystems.com/science-lab/fast-high-quality-vitrification-with-the-em-ice-high-pressure-freezer/ Thu, 08 Apr 2021 11:57:00 +0000 https://www.leica-microsystems.com/71639 CLEM Targeting Active Recycling Nuclear Pore Complexes using Cryo Confocal Microscopy In this article, how cryo light microscopy and, in particular cryo confocal microscopy, is used to improve the reliability of cryo EM workflows is described. The quality of the EM grids and samples is assessed and the distribution of target structures analyzed. It is shown how cryo confocal 3D data is projected on SEM images for improving the reliability of retaining the structure of interest within the final FIB lamella for further investigation in the cryo TEM. https://www.leica-microsystems.com/science-lab/targeting-active-recycling-nuclear-pore-complexes-using-cryo-confocal-microscopy/ Wed, 31 Mar 2021 06:09:00 +0000 Dr. Jan De Bock https://www.leica-microsystems.com/45807 CLEM EM Sample Preparation Advancing Cell Biology with Cryo-Correlative Microscopy Correlative light and electron microscopy (CLEM) advances biological discoveries by merging different microscopes and imaging modalities to study systems in 4D. Combining fluorescence microscopy with cryo-electron tomography allows investigators to bridge knowledge gaps between cell and structural biology. CLEM technologies are rapidly advancing in order to improve biological understanding, experimental reproducibility, and automation. https://www.leica-microsystems.com/advancing-cell-biology-with-cryo-correlative-microscopy/ Thu, 28 Jan 2021 08:54:00 +0000 https://www.leica-microsystems.com/44778 EM Sample Preparation High-pressure freezing: Revealing functional mechanisms of synaptic transmission Learn more about applying optogenetic stimulation in the EM ICE and how this technology has the potential to reveal structural and functional mechanisms of synaptic transmission. Get a detailed introduction of how optogenetic stimulation is applied to intact networks in acute brains slices and organotypic brain slice cultures from mice. https://www.leica-microsystems.com/high-pressure-freezing-revealing-functional-mechanisms-of-synaptic-transmission/ Mon, 14 Dec 2020 08:16:00 +0000 https://www.leica-microsystems.com/44360 EM Sample Preparation Workflows and Instrumentation for Cryo-electron Microscopy Cryo-electron microscopy is an increasingly popular modality to study the structures of macromolecular complexes and has enabled numerous new insights in cell biology. In recent years, cryo-electron microscopy has expanded further towards in situ structural biology and has become the go-to technique for resolving structures in their native context. Similarly, freeze-fracturing and cryo-scanning electron microscopy (SEM) have grown in popularity. https://www.leica-microsystems.com/workflows-and-instrumentation-for-cryo-electron-microscopy-an-overview/ Tue, 10 Nov 2020 10:42:00 +0000 PhD Frédéric Leroux https://www.leica-microsystems.com/29779 Metallography Quality Assurance Top Issues Related to Standards for Rating Non-Metallic Inclusions in Steel Supplying components and products made of steel to users worldwide can require that a single batch be compliant with multiple steel quality standards. This user demand creates significant challenges for suppliers. The standards specify rigorous methods for rating non-metallic inclusions in steel, as the inclusions have a strong influence on quality. Rating the steel quality helps ensure that products and components meet performance and safety specifications. https://www.leica-microsystems.com/science-lab/top-issues-related-to-standards-for-rating-non-metallic-inclusions-in-steel/ Fri, 19 Jun 2020 09:58:00 +0000 PhD James DeRose, PhD David R. Barbero, Thomas Locherer https://www.leica-microsystems.com/29928 Quality Assurance Metallography Analyzing non-metallic inclusions in steel Oftentimes we find ourselves caught up in tedious analyses by reticle and comparison chart, time-consuming double-evaluation according to several standards or subjective inspection results with a bias from different users. In this webinar, Dr. Nicol Ecke will talk about the advantages of automated analysis of non-metallic inclusions using LAS X Steel Expert*. Learn how this will help you get the reliable, unbiased and standard-compliant results you are aiming for faster and easier than ever before. https://www.leica-microsystems.com/science-lab/analyzing-non-metallic-inclusions-in-steel/ Tue, 02 Jun 2020 11:17:00 +0000 https://www.leica-microsystems.com/29634 EM Sample Preparation CLEM Introduction to Ion Beam Etching with the EM TIC 3X In this article you can learn how to optimize the preparation quality of your samples by using the ion beam etching method with the EM TIC 3X ion beam milling machine. A short introduction of the instrument explains how the flexible setup of the EM TIC 3X gives you the opportunity to prepare samples for various investigation areas. https://www.leica-microsystems.com/science-lab/introduction-to-ion-beam-etching-with-the-em-tic-3x/ Mon, 11 May 2020 13:15:00 +0000 https://www.leica-microsystems.com/19946 Metallography Quality Assurance Rate the Quality of Your Steel: Free Webinar and Report This webinar and report describe optimal microscopy solutions for rating steel quality in terms of non-metallic inclusions and reviews the various international and regional standards concerning rigorous quality assessment methods, e.g., EN 10247, ASTM E45, DIN 50602, and ISO 4967. https://www.leica-microsystems.com/science-lab/rate-the-quality-of-your-steel-free-webinar-and-report/ Tue, 28 Apr 2020 12:19:00 +0000 Dionis Diez, PhD James DeRose, Thomas Locherer https://www.leica-microsystems.com/20212 EM Sample Preparation Metallography Quality Assurance Stereo Microscopy 3-Dimensional Imaging of Macroscopic Defects in Aluminum Alloys The investigation of macroscale defects in aluminum (Al) alloys with a rapid 3-dimensional (3D) imaging approach is described in this report. Aluminum (Al) alloys play an important role in the production of aircraft and vehicles, as well as products in other industries. Defects present in the Al alloy used for the production of aircraft, vehicles, or other products can have a significant effect on their quality, performance, and lifetime. https://www.leica-microsystems.com/science-lab/3-dimensional-imaging-of-macroscopic-defects-in-aluminum-alloys/ Thu, 17 Oct 2019 07:00:00 +0000 PhD Wolfgang Grünewald, PhD James DeRose https://www.leica-microsystems.com/26685 EM Sample Preparation Studying the Microstructure of Natural Polymers in Fine Detail The potential of cryogenic broad ion beam milling used in combination with scanning electron microscopy (cryo-BIB-SEM) for imaging and analyzing the microstructure of cryogenically stabilized soft polymers is assessed in this report. Results from the examination of delicate, natural polymers, such as tomato skin and wood, with cryo-BIB-SEM are shown. The polymer surface morphology and various microstructural features were analyzed. https://www.leica-microsystems.com/science-lab/studying-the-microstructure-of-natural-polymers-in-fine-detail/ Wed, 16 Oct 2019 10:30:00 +0000 PhD Joyce Schmatz, PhD James DeRose, PhD David R. Barbero, M.Sc. Martin Nopens https://www.leica-microsystems.com/14345 EM Sample Preparation Introduction to Ultramicrotomy When studying samples, to visualize their fine structure with nanometer scale resolution, most often electron microscopy is used. There are 2 types: scanning electron microscopy (SEM) which images the sample surface or transmission electron microscopy (TEM) which requires a very thin, electron-transparent sample. Thus, to image the fine structure inside a sample using electron microscopy, the solution is to make very thin sections of it. https://www.leica-microsystems.com/science-lab/brief-introduction-to-ultramicrotomy/ Tue, 17 Sep 2019 09:06:00 +0000 Robert Ranner, PhD James DeRose https://www.leica-microsystems.com/27911 CLEM Improve Cryo Electron Tomography Workflow Leica Microsystems and Thermo Fisher Scientific have collaborated to create a fully integrated cryo-tomography workflow that responds to these research needs: Reveal cellular mechanisms at subnanometer resolution in 3D applying the fully integrated cryo electron tomography workflow https://www.leica-microsystems.com/science-lab/improve-cryo-electron-tomography-workflow/ Tue, 03 Sep 2019 09:50:00 +0000 https://www.leica-microsystems.com/26267 Fluorescence Microscopy Stereo Microscopy Improved tree-ring visualization using autofluorescence Autofluorescence can be a very practical tool for the visualization of growth rings in wood, especially when these structures are hard to distinguish, like in tropical trees. Tree rings are largely used for the investigation of growth rate, forest dynamics and management, paleoclimate and climate change, among other applications. https://www.leica-microsystems.com/science-lab/improved-tree-ring-visualization-using-autofluorescence/ Tue, 23 Jul 2019 16:00:00 +0000 Ph.D. Piero Bagnaresi, PhD James DeRose https://www.leica-microsystems.com/26264 Super-Resolution The Guide to STED Sample Preparation This guide is intended to help users optimize sample preparation for stimulated emission depletion (STED) nanoscopy, specifically when using the TCS SP8 STED 3X nanoscope from Leica Microsystems. It gives an overview of fluorescent labels used for single color STED imaging and a ranking of their performance. Fluorescent label combinations for dual and triple color STED imaging that minimize cross-talk during detection are recommended. There is a discussion of considerations for immunofluorescence labeling and a detailed protocol to obtain high quality images, with a high signal/noise (S/N) ratio, of interesting structures in a specimen. Important details for sample mounting and substrates that enable optimal imaging, minimizing aberrations and autofluorescence due to the mounting medium, are reviewed. Finally, for STED imaging of live-cells, the most appropriate fluorescent labels are mentioned, both fluorescent proteins (FPs) and organic fluorophores which give the best performance. https://www.leica-microsystems.com/science-lab/the-guide-to-sted-sample-preparation/ Mon, 22 Jul 2019 17:00:00 +0000 https://www.leica-microsystems.com/24656 Light Sheet Microscopy Using Mounting Frames for DLS Sample Preparation Sample handling is an important topic in the context of Light Sheet Microscopy. The TCS SP8 DLS integrates Light Sheet technology into an inverted confocal platform and can hence make use of general principles concerning sample mounting and xy-stage functionality. We describe a set of mounting frames that allow preparation of a higher number of samples, also in the context of potentially harmful mounting media like BABB. https://www.leica-microsystems.com/science-lab/using-mounting-frames-for-dls-sample-preparation/ Sun, 10 Feb 2019 23:00:00 +0000 Simone Diehl, Dr. Petra Haas https://www.leica-microsystems.com/20488 Light Sheet Microscopy Using a Rotation Device for DLS Sample Mounting The TCS SP8 DLS from Leica Microsystems is an innovative concept to integrate the Light Sheet Microscopy technology into the confocal microscope. Due to its unique optical architecture samples can be mounted on standard glass bottom petri dishes and require only little adaptations when compared to conventional mounting procedures, as for confocal imaging. https://www.leica-microsystems.com/science-lab/using-a-rotation-device-for-dls-sample-mounting/ Tue, 22 Jan 2019 23:00:00 +0000 Dr. Petra Haas, Elena Remacha Motta, Laia Ortiz Lopez, Dr. Bruno Cossermelli Vellutini, Dr. Pavel Tomancak, Dr. Emmanuel Reynaud https://www.leica-microsystems.com/20642 EM Sample Preparation High Resolution Array Tomography with Automated Serial Sectioning The optimization of high resolution, 3-dimensional (3D), sub-cellular structure analysis with array tomography using an automated serial sectioning solution, achieving a high section density on the carrier substrate, is described in this report. https://www.leica-microsystems.com/science-lab/high-resolution-array-tomography-with-automated-serial-sectioning/ Sun, 14 Oct 2018 22:00:00 +0000 Robert Ranner, PhD James DeRose https://www.leica-microsystems.com/20352 EM Sample Preparation Macroscale to Nanoscale Pore Analysis of Shale and Carbonate Rocks Physical porosity in rocks, like shale and carbonate, has a large effect on the their storage capacity. The pore geometries also affect their permeability. Imaging the visible pore space provides insights into the physical pore space, pore geometries, and the associated mineral and organic matter phases relevant for storage and transport. For example, organic-matter porosity is an important property of mature organic-rich shales providing storage capacity for liquids and gases, like hydrocarbons. Imaging the microstructure is the only technique which delivers data giving direct insight into the organic matter porosity. However, imaging porosity in fine-grained shale as well as carbonate rocks in a representative way at high resolution is challenging. https://www.leica-microsystems.com/science-lab/macroscale-to-nanoscale-pore-analysis-of-shale-and-carbonate-rocks/ Tue, 04 Sep 2018 22:00:00 +0000 PhD Jop Klaver, PhD Joyce Schmatz, MSc Mingze Jiang, PhD James DeRose https://www.leica-microsystems.com/19710 Light Sheet Microscopy DLS Sample Preparation: Using U-Shaped Glass Capillaries for Sample Mounting The TCS SP8 DLS microscope system from Leica Microsystems is an innovative concept which integrates the Light Sheet Microscopy technology into the SP8 confocal platform. Due to its unique optical architecture, samples can be mounted on standard glass bottom petri dishes and require little or no adaptation when compared to conventional mounting procedures. Here, we present a convenient way to prepare specimens quickly for light sheet imaging with the TCS SP8 DLS system. https://www.leica-microsystems.com/science-lab/dls-sample-preparation-using-u-shaped-glass-capillaries-for-sample-mounting/ Wed, 06 Dec 2017 08:45:00 +0000 Dr. Petra Haas https://www.leica-microsystems.com/18094 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/micro-ct-of-insect-larva-protocol/ Tue, 14 Mar 2017 07:14:00 +0000 PhD Elisabeth Lipke, PhD Peter Michalik https://www.leica-microsystems.com/18221 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/contrast-enhancement-of-polycrystalline-metals-sample-preparation-for-sem/ Wed, 01 Feb 2017 10:33:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/19182 EM Sample Preparation Variety of RNAs in Peripheral Blood Cells, Plasma, and Plasma Fractions Human peripheral blood contains RNA in cells and in extracellular membrane vesicles, microvesicles and exosomes, as well as in cell-free ribonucleoproteins. Circulating mRNAs and noncoding RNAs, being internalized, possess the ability to modulate vital processes in recipient cells. In this study, with SOLiD sequencing technology, we performed identification, classification, and quantification of RNAs from blood fractions: cells, plasma, plasma vesicles pelleted at 16,000 https://www.leica-microsystems.com/science-lab/variety-of-rnas-in-peripheral-blood-cells-plasma-and-plasma-fractions/ Mon, 30 Jan 2017 10:22:00 +0000 https://www.leica-microsystems.com/19176 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/drosophila-larvae-sample-preparation-for-cryo-sem/ Mon, 30 Jan 2017 09:50:00 +0000 Dr. Andres Kaech, Prof. Damian Brunner https://www.leica-microsystems.com/19179 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/giardia-lamblia-sample-preparation-for-cryo-sem/ Mon, 30 Jan 2017 09:35:00 +0000 Dr. Andres Kaech, Joe Paulin Zumthor https://www.leica-microsystems.com/18156 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-cadmiumsulphide-cds-for-cathodoluminescence/ Tue, 24 Jan 2017 18:12:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18207 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/picea-abies-l-karst-sample-preparation-for-tem/ Mon, 23 Jan 2017 19:24:00 +0000 Prof. Bernd Zechmann, Prof. Günther Zellnig https://www.leica-microsystems.com/18196 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-copper-for-electron-backscattered-diffraction-ebsd/ Thu, 19 Jan 2017 16:37:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18247 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/porous-ceramics-sample-preparation-for-sem/ Tue, 17 Jan 2017 07:32:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18319 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/maple-acer-saccharum-leaves-high-pressure-freezing-and-freeze-substitution-for-tem/ Mon, 16 Jan 2017 17:08:00 +0000 Dr. Kim Rensing https://www.leica-microsystems.com/18249 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/removal-of-surface-layers-sample-preparation-for-sem-and-tem/ Wed, 11 Jan 2017 10:10:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18354 EM Sample Preparation Cross Section of Solar Cells Application Note for Leica EM TIC020, Leica EM TIC 3X - Cross section of a complete solar cell. https://www.leica-microsystems.com/science-lab/cross-section-of-solar-cells/ Fri, 23 Dec 2016 06:47:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18362 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-a-multilayer-system-preparation-of-a-perfect-sample-surface-for-ebsd/ Thu, 22 Dec 2016 20:48:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18317 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/high-pressure-freezing-and-freeze-substitution-of-hep-2-cells-infected-with-chlamydia-pneumoniae/ Tue, 20 Dec 2016 11:03:00 +0000 Dr. Andres Kaech https://www.leica-microsystems.com/18243 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/multilayer-systems-with-widely-different-sputter-rates-sample-preparation-for-tem/ Fri, 16 Dec 2016 16:57:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18225 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/in-containing-compound-semiconductors-sample-preparation-for-tem/ Mon, 12 Dec 2016 10:13:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/19003 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/carbon-coating-for-polymeric-materials/ Mon, 05 Dec 2016 09:22:00 +0000 PhD Frédéric Leroux https://www.leica-microsystems.com/18983 EM Sample Preparation High-Resolution Carbon Coating: How much Carbon is too much? Application Note for Leica EM ACE600 - Carbon support films are routinely used for high resolution TEM. Thickness is one of the main criteria to assess its usefulness for a particular experiment. Within that respect graphene (oxide) layers are frequently used. However, charge dissipation and mechanical stability towards high probe currents and high voltages, including long term acquisition protocols are equally important. https://www.leica-microsystems.com/science-lab/high-resolution-carbon-coating-how-much-carbon-is-too-much/ Wed, 23 Nov 2016 11:56:00 +0000 PhD Frédéric Leroux https://www.leica-microsystems.com/18976 EM Sample Preparation Each Atom Counts: Protect Your Samples Prior to FIB Processing Application Note for Leica EM ACE600 - Focused ion beam (FIB) technology has become an indispensable tool for site-specific TEM sample preparation. It allows to extract electron transparent specimens with nanometer precision using a focused Ga+ ion beam. https://www.leica-microsystems.com/science-lab/each-atom-counts-protect-your-samples-prior-to-fib-processing/ Wed, 23 Nov 2016 11:43:00 +0000 PhD Frédéric Leroux https://www.leica-microsystems.com/18312 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/commercially-available-hand-creams-sample-preparation-for-cryo-sem/ Fri, 18 Nov 2016 17:22:00 +0000 Dr. Kim Rensing https://www.leica-microsystems.com/18255 EM Sample Preparation "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. https://www.leica-microsystems.com/science-lab/shallow-trench-isolation-structures-sample-preparation-for-tem/ Fri, 18 Nov 2016 16:50:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18110 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/tobacco-leaf-critical-point-drying-protocol-for-sem/ Fri, 18 Nov 2016 16:21:00 +0000 Dr. Martin W. Goldberg, M.Sc. Christine Richardson https://www.leica-microsystems.com/18245 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/paper-samples-sample-preparation-for-sem/ Wed, 16 Nov 2016 14:01:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18241 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/metal-films-and-sheets-sample-preparation-for-tem/ Wed, 16 Nov 2016 13:32:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18106 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/nematode-e-dianae-critical-point-drying-protocol-for-sem/ Wed, 16 Nov 2016 11:05:00 +0000 Mag. Nikolaus Leisch https://www.leica-microsystems.com/18940 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/visualization-of-membrane-dynamics-with-millisecond-temporal-resolution/ Mon, 07 Nov 2016 10:53:00 +0000 PhD Shigeki Watanabe https://www.leica-microsystems.com/18235 EM Sample Preparation 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". https://www.leica-microsystems.com/science-lab/contrast-enhancement-of-polished-cross-sections-of-semiconductor-structures-sample-preparation-for-sem/ Fri, 28 Oct 2016 12:08:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18806 Super-Resolution Practical Guide for Excellent GSDIM Super-Resolution Images Do you know that most protists and bacteria lack in one feature that each of our body cell has? Our cells are touch and communicate with one another. They send and receive a variety of signals that coordinate their behavior to act together as a functional multicellular organism. Exploring the way of cellular communication and the ways how the cell surface interacts to organize tissues and body structures is of great interest. Kees Jalink and his team of scientists at the Netherlands Cancer Institute (NKI) in Amsterdam obtained new scientific insights into the molecular architecture of hemidesmosomes, cytoskeletal components, cell surface receptors and vesicular proteins with the help of Ground-State-Depletion (GSD)/ dSTORM microscopy. In this interview, Kees Jalink comments on their developments in imaging chambers, buffer conditions and image analysis to get the perfect super resolution image. https://www.leica-microsystems.com/science-lab/practical-guide-for-excellent-gsdim-super-resolution-images/ Wed, 26 Oct 2016 06:58:00 +0000 PhD Kees Jalink, PhD Tamara Straube, MSc Leila Nahidiazar, MSc Daniela Leyton Puig https://www.leica-microsystems.com/18142 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/wall-cress-pod-protocol-critical-point-drying-of-arabidopsis-thaliana-for-sem/ Tue, 25 Oct 2016 08:00:00 +0000 Dr. Chen LiYu https://www.leica-microsystems.com/18257 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/surface-modification-of-znag-sample-sample-preparation-for-sem/ Mon, 17 Oct 2016 08:16:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18364 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-nicu-on-steel-for-ebsd/ Fri, 14 Oct 2016 09:45:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18096 EM Sample Preparation 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. https://www.leica-microsystems.com/science-lab/bacteria-protocol-critical-point-drying-of-e-coli-for-sem/ Thu, 13 Oct 2016 10:04:00 +0000 Dr. W. H. Mueller https://www.leica-microsystems.com/18178 EM Sample Preparation Cross Sectioning of Painted Concrete Application Note for Leica EM TIC 3X - Ion beam slope cutting is a method that can achieve flat cuts of material combinations consisting of hard and soft materials. Here, cross sectioning of painted concrete is describes in order to visualize the interface between paint and concrete. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-painted-concrete/ Mon, 10 Oct 2016 20:39:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18104 EM Sample Preparation Clawed Frog Nuclear Envelope Protocol Application Note for Leica EM CPD300 - Critical point drying of nuclear pores from clawed frog oocytes with subsequent chromium coating and SEM analysis. Silicon chips containing the samples were placed into the filter discs and porous pots holder. https://www.leica-microsystems.com/science-lab/clawed-frog-nuclear-envelope-protocol/ Thu, 06 Oct 2016 16:00:00 +0000 Dr. Martin W. Goldberg, M.Sc. Christine Richardson https://www.leica-microsystems.com/18092 EM Sample Preparation Micro-Computed Tomography (micro-CT) of Insect Brain Protocol Application Note for Leica EM CPD300 - Critical point drying of the blow fly with subsequent X-ray micro-computed tomography (micro-CT) to detect neuroanatomical features. https://www.leica-microsystems.com/science-lab/micro-computed-tomography-micro-ct-of-insect-brain-protocol/ Thu, 06 Oct 2016 14:41:00 +0000 PhD Elisabeth Lipke, PhD Peter Michalik https://www.leica-microsystems.com/18259 EM Sample Preparation Thin Metal Foils with Coatings - Sample Preparation for SEM Application Note for Leica EM RES102 - Thin foils are mostly unstable because of their thickness of a few microns. This makes it difficult to do slope cutting without any protection of the sample. A common realisation to protect the sample surface is by sticking a cover glass on top of the sample. Another issue is cutting the foils before ion milling. The sample edge should be flat and sharp without any broken areas. A razor blade is mostly the best solution. A protected sample can salso be sawed with a wire saw. https://www.leica-microsystems.com/science-lab/thin-metal-foils-with-coatings-sample-preparation-for-sem/ Wed, 05 Oct 2016 07:16:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18818 EM Sample Preparation Ultra-thin Carbon Support Films for Improved STEM-EELS Analysis of Nanoparticles Application Note for Leica EM ACE600 - Recent developments in aberration corrected transmission electron microscopes as well as further improvements in monochromaters and spectrometers have pushed the attainable energy resolution for Electron energy loss spectroscopy (EELS) to 100 meV and beyond. STEM-EELS of individual nanomaterials can be challenging due the necessity of a support film. https://www.leica-microsystems.com/science-lab/ultra-thin-carbon-support-films-for-improved-stem-eels-analysis-of-nanoparticles/ Wed, 21 Sep 2016 11:27:00 +0000 PhD Frédéric Leroux https://www.leica-microsystems.com/18820 EM Sample Preparation Ways to Reveal More from your Samples: Ultra-Thin Carbon Films Application Note for Leica EM ACE600 - Much of the battle involved in obtaining good transmission electron microscopy data is in the specimen preparation itself. Even though some nanomaterials are already electron transparent (e.g. nanoparticles and proteins) and often do not require further thinning procedures, they need to be dispersed onto thin support films. https://www.leica-microsystems.com/science-lab/ways-to-reveal-more-from-your-samples-ultra-thin-carbon-films/ Wed, 21 Sep 2016 11:23:00 +0000 PhD Frédéric Leroux, Jan de Weert https://www.leica-microsystems.com/18801 Super-Resolution Fluorescence Microscopy Measuring the 3D STED-PSF with a new Type of Fluorescent Beads A new type of fluorescent bead is presented by GATTAquant. These beads, called GATTA-Beads, are characterized by a small diameter (23 nm), high intensity and size uniformity. In combination with state-of the-art STED microscopes such as the Leica TCS SP8 STED 3X and high-end image restoration methods available in the Huygens Software, it is shown that these new beads can be used for accurate STED PSF characterization in 3D. Furthermore, it is shown that the measured 3D STED-PSF can be used to improve image restoration quality in combination with STED deconvolution methods available in the Huygens Software. https://www.leica-microsystems.com/science-lab/measuring-the-3d-sted-psf-with-a-new-type-of-fluorescent-beads/ Wed, 21 Sep 2016 06:54:00 +0000 PhD Jürgen J. Schmied, MSc Remko Dijkstra, Ph.D. Max B. Scheible, Ph.D. Giulia M. R. De Luca, PhD Jochen J. Sieber https://www.leica-microsystems.com/18192 EM Sample Preparation Cross Sectioning of a Superconductive Wire Application Note for Leica EM TIC 3X - Purpose: The shape of the wire is difficult for ion beam slope cutting. Goal: Cross sectional preparation to see the structure of the wire. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-a-superconductive-wire/ Tue, 20 Sep 2016 16:41:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18691 Stereo Microscopy Confocal Microscopy Fluorescence Microscopy Work Efficiently in Developmental Biology with Stereo and Confocal Microscopy: C. elegans For scientists, technicians, and teachers working with the worm C. elegans in the research lab or classroom, this report is intended to give useful information to help improve their daly work. The aim is to make the work steps of worm picking, transgenesis, RNA interference, screening, and functional imaging efficient. It also details the various possibilities for equipping a research worm lab or biology classroom/teaching lab explaining worm methods. https://www.leica-microsystems.com/science-lab/work-efficiently-in-developmental-biology-with-stereo-and-confocal-microscopy-c-elegans/ Mon, 19 Sep 2016 06:09:00 +0000 PhD James DeRose, PhD Heinrich Bürgers, PhD Martin Gamerdinger https://www.leica-microsystems.com/18149 EM Sample Preparation Structural Study of C. elegans Application Note for Leica EM ICE, Leica EM AFS2 - Wildtype L4 stage C. elegans (N2 strain) were placed in the 100 μm deep side of Lecithin-coated (see detailed protocol*) type A 3 mm Cu/Au carriers (Leica) with extracellular filler containing 1% (w/v) Agarose type IX and 2% (w/v) Bovine Serum Albumin in bacteria medium (see preparation details**) and sandwiched with the flat side of Lecithin-coated type B 3 mm Cu/Au carriers (Leica). Samples were frozen in a high-pressure freezer (Leica EM ICE). https://www.leica-microsystems.com/science-lab/structural-study-of-c-elegans/ Fri, 16 Sep 2016 08:15:00 +0000 E. G. van Donselaar, Dr. Martin Harterink, Drs. C. E. M. Vocking, Dr. Rob Mesman https://www.leica-microsystems.com/18344 EM Sample Preparation Cross Section of an Aluminium Sample for Electron Backscattered Diffraction (EBSD) Application Note for Leica EM TIC020, Leica EM TIC 3X - Electron backscattered diffraction (EBSD) is for example used to examine the crystallographic orientation of material. The sample preparation for such samples is sometimes very tricky as the depth of information is just few nm (~20nm or less). That means the sample surface must be flat and free of preparation artefacts. Mechanical polishing leads mostly to sample surfaces damages. https://www.leica-microsystems.com/science-lab/cross-section-of-an-aluminium-sample-for-electron-backscattered-diffraction-ebsd/ Thu, 15 Sep 2016 07:54:00 +0000 Robert Ranner https://www.leica-microsystems.com/18301 EM Sample Preparation Freeze-Fracture Replication of Pyramidal Cells Application Note for Leica EM HPM100 - Frozen samples (90 μm thick slices frozen by HPM100) were inserted into a double replica table and then fractured into two pieces at –130°C (after insertion of the tissue into BAF 060 the samples should be left in the chamber for 20 min to reach the –130°C). https://www.leica-microsystems.com/science-lab/freeze-fracture-replication-of-pyramidal-cells/ Thu, 08 Sep 2016 16:23:00 +0000 Akos Kulik https://www.leica-microsystems.com/18239 EM Sample Preparation Ion Beam Polishing of Sample Surfaces - Sample Preparation for SEM Application Note for Leica EM RES102 - Ion milling can be used to reduce the roughness of sample surfaces. Small angles less than 6° with respect to the sample surface are necessary. The high voltage depends on the material to be prepared. The reason for the levelling effect is the different milling angle of flat and rough surface areas. The milling rate is lower for small angles. The rough surface area will be faster milled. Ion polishing is often the final step of sample preparation. The prerequisite is a perfect mechanical prepreparation as samples with deep surface scratches cannot be ion polished. Soft materials usually have a smeared sample surface after mechanical polishing. It is necessary to remove this smeared material before ion polishing. Otherwise the above mentioned polishing effect does not work. https://www.leica-microsystems.com/science-lab/ion-beam-polishing-of-sample-surfaces-sample-preparation-for-sem/ Tue, 06 Sep 2016 11:11:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18215 EM Sample Preparation Cleaning of Smeared Sample Surfaces - Sample Preparation for SEM Application Note for Leica EM RES102 - Mechanical polishing of soft materials or hard / soft material combinations is tricky. The mechanical polishing process leads very often to smearing of the soft material. The smeared material covers the surface and fills small pores or holes. Grain structures, interfaces and other structural details can be masked. An additional ion milling step with milling angles between 10° C and 15° C with respect to the sample surface can remove or reduce the contamination. https://www.leica-microsystems.com/science-lab/cleaning-of-smeared-sample-surfaces-sample-preparation-for-sem/ Fri, 02 Sep 2016 06:49:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18090 EM Sample Preparation Micro-Computed Tomography (micro-CT) of Book Scorpion Musculature Application Note for Leica EM CPD300 - Critical point drying of book scorpion with subsequent X-ray micro-computed tomography (micro-CT) to detect anatomical features with special regard to the musculature. https://www.leica-microsystems.com/science-lab/micro-computed-tomography-micro-ct-of-book-scorpion-musculature/ Thu, 01 Sep 2016 08:39:00 +0000 PhD Elisabeth Lipke, PhD Peter Michalik https://www.leica-microsystems.com/18146 EM Sample Preparation Cultured Rat Hippocampal Neurons Application Note for Leica EM ICE - Rat Hippocampal neurons, cultured on 50 μm thick Aclar (Aclar embedding film, EMS) for 19 days, were frozen in the 100 μm deep side of lecithin coated (detailed protocol Appendix I) type A 3 mm Cu/Au carriers (Leica) and sandwiched with the flat side of lecithin coated type B 3 mm Cu/Au carriers (Leica). No additional filler was used, only cell culture medium with the addition of Hepes buffer pH 7.2 to a final concentration of 25 mM. Samples were frozen in a high-pressure freezer (Leica EM ICE). https://www.leica-microsystems.com/science-lab/cultured-rat-hippocampal-neurons/ Mon, 29 Aug 2016 07:12:00 +0000 E. G. van Donselaar, Dr. Martin Harterink, Drs. C. E. M. Vocking, Dr. W. H. Mueller, Prof. Dr. C. C. Hoogenraad https://www.leica-microsystems.com/18000 EM Sample Preparation Epoxy Resin Embedding of Animal and Human Tissues for Pathological Diagnosis and Research Application Note for Leica EM AMW - The tissues were fixed in the modified Karnovsky fixative generally by immersion overnight (at minimum 4h at room temperature). Then pieces of approx. 1mm3 were cut with a sharp razor blade and processed for embedding in the Leica EM AMW Microwave Tissue Processor. https://www.leica-microsystems.com/science-lab/epoxy-resin-embedding-of-animal-and-human-tissues-for-pathological-diagnosis-and-research/ Fri, 19 Aug 2016 08:34:00 +0000 Dr. Josef A. Schroeder https://www.leica-microsystems.com/18209 EM Sample Preparation Ceramics - Sample Preparation for TEM Application Note for Leica EM RES102 - Ceramic samples are mostly very brittle, and are therefore very difficult to thin mechanically to a low starting thickness for ion beam milling. The ion beam milling of insulators often leads to static charging of the surface of the sample. This, in turn, reduces the sputter rate. When using the Ti standard holder (standard TEM holder), however, sufficient secondary electrons are created by the ion beam also falling on the sample holder to largely compensate for the static charging. https://www.leica-microsystems.com/science-lab/ceramics-sample-preparation-for-tem/ Mon, 15 Aug 2016 18:46:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18295 EM Sample Preparation Cryo-Electron Microscopy of Vitreous Sections (CEMOVIS) of Yeast Application Note for Leica EM HPM100 - The sections are of yeast frozen with a Leica HPM100 high pressure freezer in the copper tube system, the cell paste was mixed with a pH 6.5 MES/dextran buffer so that a final MES concentration of 50mM and a dextran concentration of 20% was achieved. https://www.leica-microsystems.com/science-lab/cryo-electron-microscopy-of-vitreous-sections-cemovis-of-yeast/ Fri, 12 Aug 2016 14:51:00 +0000 Dr. Jonathan O'Driscoll, Dr. Daniel Kofi Clare, Prof. Helen Saibil https://www.leica-microsystems.com/18144 EM Sample Preparation Wrinkled Giant Hyssop Leaf Protocol Application Note for Leica EM CPD300 - Critical point drying of wrinkled giant hyssop leaf with subsequent gold coating and SEM analysis. https://www.leica-microsystems.com/science-lab/wrinkled-giant-hyssop-leaf-protocol/ Thu, 11 Aug 2016 17:04:00 +0000 Dr. Guo JianSheng https://www.leica-microsystems.com/18100 EM Sample Preparation Black Mould Protocol Application Note for Leica EM CPD300 - Critical point drying of Black mould (Aspergilus niger) with subsequent platinum / palladium coating and SEM analysis to detect conidiospores. Sample was inserted into a filter disc (Pore size: 16 - 40 μm) and placed into the filter discs and porous pots holder. https://www.leica-microsystems.com/science-lab/black-mould-protocol/ Tue, 09 Aug 2016 12:01:00 +0000 Dr. W. H. Mueller https://www.leica-microsystems.com/18151 EM Sample Preparation Cross Sectioning of Alumina Application Note for Leica EM TIC 3X - Alumina is very difficult to handle and almost impossible to prepare with conventional methods. Ion beam slope cutting is a method that can achieve cross sections of material combinations consisting of hard and soft components. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-alumina/ Mon, 08 Aug 2016 07:48:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/17998 EM Sample Preparation Arabidopsis thaliana(L.) Accession Col Application Note for Leica EM AMW - Plants were grown in growth chambers under defined conditions. After stratification for 4 days at 4°C seeds were grown in pots with soil with 9/15 hours day/night photoperiod. Day and night temperatures were 22°C and 18°C, respectively, the relative humidity was 60% and the plants were kept at 100% relative soil water content. Light intensity varied between 110 and 140 μmol m-2 s-1. https://www.leica-microsystems.com/science-lab/arabidopsis-thalianal-accession-col/ Mon, 01 Aug 2016 13:03:00 +0000 Prof. Bernd Zechmann, Prof. Günther Zellnig https://www.leica-microsystems.com/18190 EM Sample Preparation Cross Sectioning of CuSn Connector of a Solar Cell Application Note for Leica EM TIC 3X - Ion beam slope cutting is a method that can achieve flat cross sections of soft materials or material combinations consisting of hard and soft components. The CuSn connector is very soft. Mechanical polishing leads to smearing. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-cusn-connector-of-a-solar-cell/ Fri, 29 Jul 2016 11:56:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/17987 EM Sample Preparation Immuno - Electron Microscopy of High Pressure Frozen and Freeze Substituted Mouse Heart Application Note for Leica EM AFS2 - Mouse heart tissue from wild-type (WT) and αT -catenin KO (KO) mice was excised, immersed in 20% (w/v) BSA and frozen immediately in a high-pressure freezer Leica EM PACT. Freeze subs­titution was carried out using a Leica EM AFS in dry acetone containing 2% ddH2O and 0.1% glutaraldehyde over a 4-days period. https://www.leica-microsystems.com/science-lab/immuno-electron-microscopy-of-high-pressure-frozen-and-freeze-substituted-mouse-heart/ Wed, 27 Jul 2016 15:35:00 +0000 Riet de Rycke https://www.leica-microsystems.com/17979 EM Sample Preparation Cell Envelope Structure of a Gram-negative Thermotolerant y-Proteobacterium Acidithiobacillus sp., Strain HV2/2 and its Interaction with Pyrite Application Note for Leica EM AFS2 - Cells of the the moderately thermophilic Acidithiobacillus sp.strain HV2/2, were centrifuged at 20,0009xg and then cryo-immobilized by high-pressure freezing on gold carriers. https://www.leica-microsystems.com/science-lab/cell-envelope-structure-of-a-gram-negative-thermotolerant-y-proteobacterium-acidithiobacillus-sp-strain-hv22-and-its-interaction-with-pyrite/ Mon, 25 Jul 2016 10:12:00 +0000 Dr. Andreas Klingl https://www.leica-microsystems.com/18088 Super-Resolution Translation Microscopy (TRAM) for Super-Resolution Imaging Super-resolution microscopy is transforming our understanding of biology but accessibility is limited by its technical complexity, high costs and the requirement for bespoke sample preparation. We present a novel, simple and multi-color super-resolution microscopy technique, called translation microscopy (TRAM), in which a super-resolution image is restored from multiple diffraction-limited resolution observations using a conventional microscope whilst translating the sample in the image plane. https://www.leica-microsystems.com/science-lab/translation-microscopy-tram-for-super-resolution-imaging/ Fri, 22 Jul 2016 17:51:00 +0000 https://www.leica-microsystems.com/18049 EM Sample Preparation Neuroscience Symmetric Synapse - Clathrin Coated Endocytosis Pit in the Postsynaptic Dendrite Application Note for Leica EM ICE - WT hippocampal neurons were plated at a density of 80,000 cells/cm2 on 6 mm sapphire disks for 14 days. Sample were frozen using a high-pressure freezer (Leica EM ICE) under a pressure of 2100bar by mounting it into a sandwich support with extracellular solution containing 15% of Ficoll 400, to assess ice crystal damage. The Cryo-fixation was achieved within milliseconds allowing simultaneous immobilization of all macromolecular components. After freezing, sam­ple was transferred into cryovials containing 1% glutaraldehyde, 1% osmium tetroxide, 1% milliQwater in anhydrous acetone and processed in an automated freeze-substitution device (Leica EM AFS2). https://www.leica-microsystems.com/science-lab/symmetric-synapse-clathrin-coated-endocytosis-pit-in-the-postsynaptic-dendrite/ Wed, 20 Jul 2016 08:56:00 +0000 Dr. Shuwen Chang https://www.leica-microsystems.com/17975 EM Sample Preparation Electron Microscopy of High Pressure Frozen and Freeze Substituted Arabidopsis Thaliana Root Tips Cells Application Note for Leica EM AFS2 - Arabidopsis thaliana roots (mutant PIN1pro:PIN1-GFP;bex5-1) were excised, immersed in 20% (w/v) BSA and frozen immediately in a high-pressure freezer. Freeze substitution was carried out using a Leica EM AFS2. https://www.leica-microsystems.com/science-lab/electron-microscopy-of-high-pressure-frozen-and-freeze-substituted-arabidopsis-thaliana-root-tips-cells/ Fri, 08 Jul 2016 10:02:00 +0000 Riet de Rycke https://www.leica-microsystems.com/18368 EM Sample Preparation Cross Sectioning of Oil Shale Rock Application Note for Leica TIC 3X - High quality sample preparation of large area to investigate the sample in the SEM. For the mechanical preparation step diamond lapping foils of 9μm subsequently 2μm and finally 0.5μm grain size were used. It took about 1.5 hours. The sample was removed from the stub with a razor blade after TXP processing and fixed onto the holder of the rotary stage of the Leica EM TIC 3X. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-oil-shale-rock/ Tue, 28 Jun 2016 13:04:00 +0000 Robert Ranner https://www.leica-microsystems.com/18284 EM Sample Preparation Cryo-SEM Imaging of Latex Paint Application Note for Leica EM VCT100, Leica EM ACE600 - A thin layer of latex paint was spread on a clean, scored, silicon chip. The sample was immediately plunge frozen in liquid ethane and transferred under LN2 to the Leica EM VCT100 loading station where it was placed in the customized sample holder. https://www.leica-microsystems.com/science-lab/cryo-sem-imaging-of-latex-paint/ Thu, 16 Jun 2016 16:50:00 +0000 Dr. Levi Felts, Dr. Kim Rensing, Chris Frethem https://www.leica-microsystems.com/18184 EM Sample Preparation Cross Sectioning of Rubber (Tire) Application Note for Leica EM TIC 3X - Ion beam slope cutting is a method that can achieve cross sections of soft materials or material combinations consisting of hard and soft components. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-rubber-tire/ Tue, 14 Jun 2016 10:30:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/18086 EM Sample Preparation Water Flea Protocol Application Note for Leica EM CPD300 - Critical point drying of water flea with subsequent gold coating and SEM-Analysis to detect finde surface structures. https://www.leica-microsystems.com/science-lab/water-flea-protocol/ Tue, 07 Jun 2016 07:06:00 +0000 Mag. Dr. Daniela Gruber https://www.leica-microsystems.com/17941 Laser Microdissection Workflows & Protocols: Plant Laser Microdissection During Leica workshops for LMD users in Brazil, hosted by the Federal University of Paraná/UFPR (UFPR) at the Centro de Energia Nuclear na Agricultura/USP (CENA), the power of laser microdissection using the Leica LMD systems was demonstrated. One special focus was on plant dissection which needs a high laser power. https://www.leica-microsystems.com/science-lab/workflows-protocols-plant-laser-microdissection/ Mon, 06 Jun 2016 05:22:00 +0000 Dr. Falk Schlaudraff, Dr. Christoph Greb https://www.leica-microsystems.com/18079 EM Sample Preparation Human Blood Cells Protocol Application Note for Leica EM CPD300 - Life Science Research. Species: Human (Homo sapiens) Critical point drying of human blood with subsequent platinum / palladium coating and SEM analysis. https://www.leica-microsystems.com/science-lab/human-blood-cells-protocol/ Tue, 24 May 2016 08:03:00 +0000 Dr. W. H. Mueller https://www.leica-microsystems.com/17971 EM Sample Preparation Cross Sectioning of Basalt Fibres Application Note for Leica EM TIC 3X - Material Research. Purpose: The fibres are embedded in a soft matrix. That makes it difficult to prepare a cross section. Goal: Cross section of the basalt fibres. https://www.leica-microsystems.com/science-lab/cross-sectioning-of-basalt-fibres/ Fri, 20 May 2016 15:31:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/17939 Laser Microdissection Workflows & Protocols: Laser Microdissection for Pathology and Cancer Research Tumor development results from mutations in our DNA. For their deeper analysis, cancer researchers have to dissect the relevant tissue areas. Here we report the reason why laser microdissection is a perfect tool for this purpose and how this was taught in the course of a workshop held in Brazil. With the Leica LMD system pure tumor material can be selected and dissected for downstream analysis to ensure 100% pure starting material without any risk of cross contamination with healthy cells. https://www.leica-microsystems.com/science-lab/workflows-protocols-laser-microdissection-for-pathology-and-cancer-research/ Wed, 04 May 2016 11:30:00 +0000 Dr. Falk Schlaudraff, Dr. Christoph Greb https://www.leica-microsystems.com/17645 EM Sample Preparation Improvement of Metallic Thin Films for HR-SEM by Using DC Magnetron Sputter Coater Preparation techniques, like several kinds of coating methods play an important role for high resolution scanning electron microscopy (HR-SEM). Nonconductive sample like biological and synthetic samples have to be prepared with a thin conductive layer to prevent charging. https://www.leica-microsystems.com/science-lab/improvement-of-metallic-thin-films-for-hr-sem-by-using-dc-magnetron-sputter-coater/ Wed, 13 Apr 2016 08:30:00 +0000 Dr. Johannes Rattenberger, DI Alexander Melischnig, DI Dr. Ilse Letofsky-Papst, DI Thomas Ganner, Ing. Hartmuth Schröttner https://www.leica-microsystems.com/17667 Live-Cell Imaging Widefield Microscopy How to do a Proper Cell Culture Quick Check In order to successfully work with mammalian cell lines, they must be grown under controlled conditions and require their own specific growth medium. In addition, to guarantee consistency their growth must be monitored at regular intervals. This article describes a typical workflow for subculturing an adherent cell line with detailed illustrations of all of the necessary steps. https://www.leica-microsystems.com/science-lab/how-to-do-a-proper-cell-culture-quick-check/ Thu, 24 Mar 2016 09:18:00 +0000 PhD Tamara Straube, Claudia Müller https://www.leica-microsystems.com/17377 EM Sample Preparation Triple-beam Ar-Ion-Milling with a Rotary Stage to Decorate Grain Boundaries and Substructures in Rock Salt Decoration of grain boundaries in polycrystalline rocks has a long tradition in Structural Geology as in a monomineralic rock the recrystallized grain size is a good indicator for the paleostress conditions. Understanding the mechanical properties of rock salt and its deformation behavior is of major importance for the prediction of long-term stability of nuclear waste repositories, and our understanding of the dynamics of salt-related sedimentary basins which host the majority of oil and gas accumulations on Earth. https://www.leica-microsystems.com/science-lab/triple-beam-ar-ion-milling-with-a-rotary-stage-to-decorate-grain-boundaries-and-substructures-in-rock-salt/ Thu, 03 Mar 2016 11:01:00 +0000 PhD Joyce Schmatz, PhD Oliver Schenk, PhD Jop Klaver, PhD Janos Urai, PhD Wolfgang Grünewald https://www.leica-microsystems.com/17358 EM Sample Preparation TEM Sample Preparation Made Easy - Prepare TEM Specimen by Broad Beam Argon Ion Milling Quantitative and analytical analysis at high spatial resolution places stringent demands on the quality of the produced TEM specimens. Pristine and high-quality samples are indispensible for atomic resolution TEM analysis. In this application note a general procedure for obtaining cross-sectional and plan-view TEM specimens using the Leica EM RES102 ion milling system is outlined. The procedure described below can be easily adapted for a large range of materials e.g. thin film materials, semiconductors, multilayered materials, ceramics, superconductors, … https://www.leica-microsystems.com/science-lab/tem-sample-preparation-made-easy-prepare-tem-specimen-by-broad-beam-argon-ion-milling/ Tue, 15 Dec 2015 12:03:00 +0000 PhD Frédéric Leroux https://www.leica-microsystems.com/16814 Laser Microdissection Neuroscience Workflows & Protocols: Connecting Microscopy and Molecular Biology in Neuroscience The main topic during this course was how to apply laser microdissection in neuroscience. Leica specialists demonstrated why laser microdissection is a suitable techniques for brain investigation as it allows to separate distinct brain layers or even to isolate individual neurons. https://www.leica-microsystems.com/science-lab/workflows-protocols-connecting-microscopy-and-molecular-biology-in-neuroscience/ Fri, 04 Dec 2015 16:01:00 +0000 Dr. Falk Schlaudraff https://www.leica-microsystems.com/16808 Laser Microdissection Workflows & Protocols: How to Isolate Individual Chromosomes with Laser Microdissection During the first Leica Workshop in Brazil, at the Centro de Energia Nuclear na Agricultura/USP (CENA), the participants learned how to prepare samples for laser microdissection (LMD) using a cryotome. Another topic was the dissection of individual chromosomes from chromosome spreads. Leica specialists held a short training session with the LMD. After this, new LMD users were able to run the system and practiced how to dissect chromosomes and collect single chromosomes for downstream analysis. https://www.leica-microsystems.com/science-lab/workflows-protocols-how-to-isolate-individual-chromosomes-with-laser-microdissection/ Thu, 03 Dec 2015 20:06:00 +0000 Dr. Falk Schlaudraff https://www.leica-microsystems.com/16804 EM Sample Preparation Electron Microscopy Sample Preparation: “The Future is Cold, Dynamic and Hybrid” In 2014, the renowned Electron Microscopy for Materials Science (EMAT) research lab at the University Antwerp, Belgium, and Leica Microsystems started a fruitful collaboration to establish a Leica Reference Site in Antwerp. This site, officially opened in July 2014, is dedicated to specimen preparation for electron microscopy in materials science with a special focus on ion beam milling and recently also on carbon coating. In this interview Prof Gustaf van Tendeloo, Director of EMAT, and Frédéric Leroux, TEM specimen preparation specialist, talk about research topics at EMAT, how the Leica reference site has evolved, and future trends for EM sample preparation. https://www.leica-microsystems.com/science-lab/electron-microscopy-sample-preparation-the-future-is-cold-dynamic-and-hybrid/ Thu, 22 Oct 2015 19:22:00 +0000 Prof. Gustaaf (Staf) Van Tendeloo, PhD Frédéric Leroux, Florence Hauger https://www.leica-microsystems.com/16165 EM Sample Preparation A Good Place for Materials Scientists and Mineralogists to prepare their EM Samples In June 2014, the Institut de Minéralogie, de Physique des Matériaux et de Cosmochimie (IMPMC) of the Université Pierre et Marie Curie in Paris became a Leica reference lab for EM Sample Preparation with equipment like the Leica EM TXP target surfacing systems and the Leica EM TIC 3X ion beam milling system. In the interview, Imène Estève, engineer at the Centre National de Recherches Scientifiques (CNRS) and head of the SEM-FIB national facility, in charge of running the lab, tells us about how this cooperation has developed. https://www.leica-microsystems.com/science-lab/a-good-place-for-materials-scientists-and-mineralogists-to-prepare-their-em-samples/ Fri, 07 Aug 2015 09:31:00 +0000 MPhys, MChem Imène Esteve, Florence Hauger, Claudia Müller https://www.leica-microsystems.com/16110 Laser Microdissection Workflows & Protocols: How to Use a Leica Laser Microdissection System and Qiagen Kits for Successful RNA Analysis Laser Microdissection (LMD) allows isolating individual cells or chromosomes and is a well established technique for sample preparation prior downstream analysis of the nucleic acid content via PCR or sequencing techniques. Here we describe the successful combination of the Leica Microsystems LMD system and Qiagen kits for purification of nucleic acids even from little amounts. https://www.leica-microsystems.com/science-lab/workflows-protocols-how-to-use-a-leica-laser-microdissection-system-and-qiagen-kits-for-successful-rna-analysis/ Fri, 10 Jul 2015 14:26:00 +0000 Dr. Falk Schlaudraff https://www.leica-microsystems.com/16010 EM Sample Preparation Critical-point Drying for the Preparation of Biological Samples for MicroCT Analysis X-ray micro-computed tomography (micro-CT) is a routinely applied non-invasive technique for the investigation of the internal anatomy and morphology of organisms. As a result of a micro-CT scan a stack of grey-scale images is generated from a series of projections taken at defined angles during sample rotation. Since several years the number of lab-based micro-CT imaging systems is constantly growing making this technique available to a broad spectrum of researchers and applications. https://www.leica-microsystems.com/science-lab/critical-point-drying-for-the-preparation-of-biological-samples-for-microct-analysis/ Mon, 06 Jul 2015 13:23:00 +0000 PhD Peter Michalik, PhD Elisabeth Lipke https://www.leica-microsystems.com/15596 EM Sample Preparation Transmission Electron Microscopy Sample Preparation Protocols for the Ultrastructural Study of Cysts of Free-living Protozoa Cysts of free-living protozoa have an impact on the ecology and epidemiology of bacteria because they may act as a transmission vector or shelter the bacteria against hash environmental conditions. Detection and localization of intracystic bacteria and examination of the en- and excystment dynamics is a major challenge because no detailed protocols for ultrastructural analysis of cysts are currently available. https://www.leica-microsystems.com/science-lab/transmission-electron-microscopy-sample-preparation-protocols-for-the-ultrastructural-study-of-cysts-of-free-living-protozoa/ Thu, 02 Jul 2015 17:25:00 +0000 https://www.leica-microsystems.com/15096 Stereo Microscopy Intravenous Microinjections of Zebrafish Larvae to Study Acute Kidney Injury We describe a technique of microinjecting the aminoglycoside, gentamicin, into 2 days post-fetilization (dpf) zebrafish larvae to induce acute kidney injury (AKI). We also describe a method for whole mount immunohistochemistry, plastic embedding and sectioning of zebrafish larvae to visualize the AKI mediated damage. https://www.leica-microsystems.com/science-lab/intravenous-microinjections-of-zebrafish-larvae-to-study-acute-kidney-injury/ Fri, 19 Jun 2015 20:26:00 +0000 https://www.leica-microsystems.com/15930 EM Sample Preparation High Quality Sample Preparation for EBSD Analysis by Broad Ion Beam Milling Electron Backscatter Diffraction technique (EBSD) is known as a "surface" technique because electron diffraction is generated within a few tens of nanometers of the sample surface. Therefore, the specimen surface should be exempt of any damages in order to produce EBSD patterns. Here, we present a successful and efficient EBSD sample polishing of two very challenging specimens prepared by broad ion beam milling. https://www.leica-microsystems.com/science-lab/high-quality-sample-preparation-for-ebsd-analysis-by-broad-ion-beam-milling/ Fri, 12 Jun 2015 20:30:00 +0000 PhD Laurie Palasse, PhD Wolfgang Grünewald https://www.leica-microsystems.com/15509 CLEM Correlative In-Resin Super-Resolution and Electron Microscopy Using Standard Fluorescent Proteins We introduce a method for correlative in-resin super-resolution fluorescence and electron microscopy (EM) of biological structures in mammalian culture cells. Cryo-fixed resin embedded samples offer superior structural preservation, performing in-resin super-resolution, however, remains a challenge. https://www.leica-microsystems.com/science-lab/correlative-in-resin-super-resolution-and-electron-microscopy-using-standard-fluorescent-proteins/ Thu, 04 Jun 2015 10:28:00 +0000 https://www.leica-microsystems.com/15227 EM Sample Preparation Preparation of Small Disk-shaped Samples for Capacitance and Pyroelectric Current Measurements Capacitance and pyroelectric current measurements are best suited to determine important dielectric properties of materials, e.g. the relative dielectric constant, εr, and the remnant electric polarisation, P, of materials, respectively. In this technical report, we demonstrate the successful use of the Leica EM TXP surface targeting system to prepare a small capacitor. https://www.leica-microsystems.com/science-lab/preparation-of-small-disk-shaped-samples-for-capacitance-and-pyroelectric-current-measurements/ Fri, 29 May 2015 14:11:00 +0000 MSc, PhD, Prof. Andrea Gauzzi https://www.leica-microsystems.com/13642 CLEM EM Sample Preparation Correlative Light and Electron Microscopy – Get Your Free CLEM e-Book for Download The urge to go deeper into the microscopic world has led researchers to combine the versatility of the light microscopy (LM) with the resolution power of the electron microscope (EM) to produce Correlative Light and Electron Microscopy (CLEM). CLEM’s most significant and powerful characteristic for cell biology research is an ability to study the same cell using two different microscopy platforms. The Essential Knowledge Briefing describes the basics of CLEM and also reviews potential pitfalls and problems as well as tricks for solving them. https://www.leica-microsystems.com/science-lab/correlative-light-and-electron-microscopy-get-your-free-clem-e-book-for-download/ Thu, 28 May 2015 16:34:00 +0000 Professor Paul Verkade https://www.leica-microsystems.com/15287 Quality Assurance Metallography Five Advantages of Inverted Over Upright Microscopes in Industrial Applications With inverted microscopes, you look at samples from below since their optics are placed under the sample, with upright microscopes you look at samples from above. Traditionally, inverted microscopes are used for life science research, because gravity makes samples sink to the bottom of a holder with aqueous solution and you don’t see a lot from above. https://www.leica-microsystems.com/science-lab/five-advantages-of-inverted-over-upright-microscopes-in-industrial-applications/ Mon, 16 Mar 2015 09:06:00 +0000 Dr. Kay Scheffler, Claudia Müller https://www.leica-microsystems.com/14828 Confocal Microscopy Laser Microdissection CLEM Live-Cell Imaging Customized Patterned Substrates for Highly Versatile Correlative Light-Scanning Electron Microscopy Correlative light electron microscopy (CLEM) combines the advantages of light and electron microscopy, thus making it possible to follow dynamic events in living cells at nanometre resolution. Various CLEM approaches and devices have been developed, each of which has its own advantages and technical challenges. We here describe our customized patterned glass substrates, which improve the feasibility of correlative fluorescence/confocal and scanning electron microscopy. https://www.leica-microsystems.com/science-lab/customized-patterned-substrates-for-highly-versatile-correlative-light-scanning-electron-microscopy/ Fri, 06 Mar 2015 17:14:00 +0000 https://www.leica-microsystems.com/15081 EM Sample Preparation CLEM New Insights into Cilia and Flagella by Cryo-EM Cilia and flagella were the first organelles to be discovered and have been studied for centuries. However, their essential role in humans and how ciliary defects cause diseases are still not well understood. Cryo-EM has recently shed new light on their inner workings and solved some long-standing mysteries, only to raise new questions on how cilia and flagella function. https://www.leica-microsystems.com/science-lab/new-insights-into-cilia-and-flagella-by-cryo-em/ Mon, 16 Feb 2015 09:25:00 +0000 PhD Thomas Heuser https://www.leica-microsystems.com/15104 EM Sample Preparation Immersion Freezing for Cryo-Transmission Electron Microscopy: Applications A well established usage case for cryo-TEM is three-dimensional reconstruction of isolated macromolecules, virus particles, or filaments. On one hand, these approaches are based on averaging of repetitive structures – either due to numerous identical molecules, repetitive patterns on a filament, or symmetries, to reduce the noise inherent to cryo-TEM. https://www.leica-microsystems.com/science-lab/immersion-freezing-for-cryo-transmission-electron-microscopy-applications/ Mon, 09 Feb 2015 14:51:00 +0000 Dr. Guenter Resch https://www.leica-microsystems.com/15102 EM Sample Preparation Immersion Freezing for Cryo-Transmission Electron Microscopy: Fundamentals The high vacuum required in a transmission electron microscope (TEM) greatly impairs the ability to study specimens naturally occurring in an aqueous phase: exposing "wet" specimens to a pressure significantly lower than the vapour pressure of water will lead to the water phase boiling off rapidly in the column, with devastating consequences for the structure of the specimen. Hence, various methods to dry specimens before inspection are employed in conventional TEM – a preparative step often associated with artifacts limiting the significance of the results. https://www.leica-microsystems.com/science-lab/immersion-freezing-for-cryo-transmission-electron-microscopy-fundamentals/ Mon, 09 Feb 2015 14:41:00 +0000 Dr. Guenter Resch https://www.leica-microsystems.com/14887 EM Sample Preparation Video Tutorials: Filling and Assembling of Different Carriers for High-Pressure Freezing High pressure freezing (HPF) is a cryo-fixation method primarily for biological samples, but also for a variety of non-biological materials. It is a technique that yields optimal preservation in many cell types and tissues or in organic and inorganic composites. Most commonly, the high pressure frozen samples are analyzed further with light or electron microscopy after appropriate processing. https://www.leica-microsystems.com/science-lab/video-tutorials-filling-and-assembling-of-different-carriers-for-high-pressure-freezing/ Fri, 05 Dec 2014 08:45:00 +0000 Dr. Cveta Tomova https://www.leica-microsystems.com/14386 EM Sample Preparation How to Clean a Coater Coating of samples is required in the field of electron microscopy to enable or improve the imaging of samples. Compared to the traditional coater design, all parts of a Leica EM ACE Coater can be individually removed and cleaned or, if special cleanliness is needed, even exchanged for spare parts. https://www.leica-microsystems.com/science-lab/how-to-clean-a-coater/ Thu, 16 Oct 2014 16:24:00 +0000 Gisela Höflinger https://www.leica-microsystems.com/14347 EM Sample Preparation Brief Introduction to Specimen Trimming Before ultrathin sectioning a sample with an ultramicrotome it has to be pre-prepared. For this pre-preparation, special attention must be paid to the sample size (size of the section), location of the sample (targeting) and accuracy of the block-face edges. This process is generally called trimming, wherein the sample is shaped mainly to a frustum of a pyramid. https://www.leica-microsystems.com/science-lab/brief-introduction-to-specimen-trimming/ Mon, 22 Sep 2014 13:56:00 +0000 Robert Ranner https://www.leica-microsystems.com/14343 EM Sample Preparation Glycerol Spraying/Platinum Low Angle Rotary Shadowing of DNA with the Leica EM ACE600 e-beam Glycerol spraying/low angle rotary shadowing (Aebi and Baschong, 2006) is a preparation technique used in biology to visualize structures yielding insufficent contrast with other techniques, due to their small diameter. This method is commonly used for specimens which include proteins with coiled coil domains or DNA. https://www.leica-microsystems.com/science-lab/glycerol-sprayingplatinum-low-angle-rotary-shadowing-of-dna-with-the-leica-em-ace600-e-beam/ Thu, 04 Sep 2014 14:33:00 +0000 Dr. Guenter Resch https://www.leica-microsystems.com/13837 EM Sample Preparation Partner Lab for EM Sample Preparation in Paris At the Institut de Minéralogie, de Physique des Matériaux et de Cosmochimie (IMPMC) of the Université Pierre et Marie Curie in Paris an new lab for EM Sample Preparation welcomes researchers of materials sciences and mineralogy: the lab has been equipped with Leica EM TXP target surfacing systems and Leica EM TIC3X ion beam systems. https://www.leica-microsystems.com/science-lab/partner-lab-for-em-sample-preparation-in-paris/ Fri, 27 Jun 2014 07:13:00 +0000 Guillaume Fiquet https://www.leica-microsystems.com/13800 EM Sample Preparation Analysis of Oil Shale as an Alternative Source of Energy For some years now, the search for alternative sources of raw materials has concentrated on oil shale deposits. As the exploitation of these raw materials is still extremely complex and expensive, analysis of oil shale samples is therefore extremely important for identifying the deposit’s potential in advance and optimizing mining methods. https://www.leica-microsystems.com/science-lab/analysis-of-oil-shale-as-an-alternative-source-of-energy/ Tue, 24 Jun 2014 08:23:00 +0000 PhD Wolfgang Grünewald https://www.leica-microsystems.com/13786 EM Sample Preparation Metallography Triple Ion Beam Cutting of Diamond/Al Composites for Interface Characterization The microstructure of new materials with heterogenic components is extremely difficult to study. As mechanical polishing often not leads to smooth surfaces, a novel ion beam cutting technique has proven to be very effective. In this article, the applicability of a novel triple ion beam (TIB) cutting technique for a clean metallographic preparation of the diamond/Al composites will be described. https://www.leica-microsystems.com/science-lab/triple-ion-beam-cutting-of-diamondal-composites-for-interface-characterization/ Sat, 21 Jun 2014 09:22:00 +0000 PhD Gang Ji, PhD Wolfgang Grünewald https://www.leica-microsystems.com/13280 EM Sample Preparation Neuroscience Capturing Cellular Dynamics with Millisecond Temporal Resolution The combination of two powerful techniques: optogenetics and high-pressure freezing now makes it possible to visualize a dynamic cellular activity with temporal resolution of 5 milliseconds. By coupling a flash of light with high-pressure freezing, the process of vesicle recycling at the synapses can now be imaged by electron microscopy. https://www.leica-microsystems.com/science-lab/capturing-cellular-dynamics-with-millisecond-temporal-resolution/ Mon, 12 May 2014 13:16:00 +0000 PhD Shigeki Watanabe, PhD Erik M. Jørgensen https://www.leica-microsystems.com/11000 Confocal Microscopy Neuroscience Live-Cell Imaging Imaging Pheromone Sensing in a Mouse Vomeronasal Acute Tissue Slice Preparation In mice, the ability to detect pheromones is principally mediated by the vomeronasal organ (VNO). Here, an acute tissue slice preparation of VNO for performing calcium imaging is described. This physiological approach allows observations of subpopulations and/or individual neurons in a living tissue and is convenient for receptor-ligand identification. https://www.leica-microsystems.com/science-lab/imaging-pheromone-sensing-in-a-mouse-vomeronasal-acute-tissue-slice-preparation/ Fri, 02 May 2014 14:24:00 +0000 https://www.leica-microsystems.com/12779 Super-Resolution Sample Preparation for GSDIM Localization Microscopy – Protocols and Tips The widefield super-resolution technique GSDIM (Ground State Depletion followed by individual molecule return) is a localization microscopy technique that is capable of resolving details as small as 20 nanometers. GSDIM is suitable for a wide range of samples. https://www.leica-microsystems.com/science-lab/sample-preparation-for-gsdim-localization-microscopy-protocols-and-tips/ Tue, 01 Apr 2014 12:40:00 +0000 Dr. Marko Lampe, Dr. Christoph Greb https://www.leica-microsystems.com/11782 EM Sample Preparation Freeze Substitution of Trypanosoma brucei Chemical fixation of biological specimens for ultrastructural investigation is a relatively slow and selective process, and therefore a common source of artifacts. Freezing, on the other hand, is an excellent method to physically fix biological specimens in their entirety and without delay; the formation of ice crystals large enough to displace cellular material and destroy structures, would be, however, a major issue. https://www.leica-microsystems.com/science-lab/freeze-substitution-of-trypanosoma-brucei/ Tue, 18 Mar 2014 17:25:00 +0000 Harald Kotisch, Dr. Katy Schmidt, Dr. Jan Leunissen, Dr. Guenter Resch https://www.leica-microsystems.com/11772 EM Sample Preparation Protein Trafficking in Cereal Seeds Seeds accumulate proteins and starch which will be broken down and mobilized upon germination. Among seeds, cereals constitute an example of a highly specialized storage tissue that constitutes up to 80% of the total seed volume (Watson 1987). The seed endomembrane system is highly specialized and seed storage proteins travel through the endomembrane system en route to the protein bodies, which are either derived from the Endoplasmic reticulum (ER) or of vacuolar origin (Muntz 1998). https://www.leica-microsystems.com/science-lab/protein-trafficking-in-cereal-seeds/ Mon, 10 Feb 2014 17:31:00 +0000 PhD Elsa Arcalis, PhD, MSc Eva Stöger https://www.leica-microsystems.com/11809 EM Sample Preparation Brief Introduction to Freeze Substitution Freeze-substitution is a process of dehydration, performed at temperatures low enough to avoid the formation of ice crystals and to circumvent the damaging effects observed after ambient-temperature dehydration. During freeze substitution the “frozen” water is dissolved by an organic solvent, which usually also contains chemical fixatives. https://www.leica-microsystems.com/science-lab/brief-introduction-to-freeze-substitution/ Tue, 04 Feb 2014 09:32:00 +0000 Riet de Rycke, Dr. Andres Kaech https://www.leica-microsystems.com/12284 Super-Resolution The Force of the Dark Side – Embedding Media for GSDIM Super-Resolution Localization Microscopy Super-resolution microscopy such as Stimulated Emission Depletion (STED) and single-molecule based techniques rely on the same principle for breaking the diffraction limit: the unwanted fluorescence signals are switched off during the image acquisition process. Consequently, Ground State Depletion followed by Individual Molecule Return (GSDIM) microscopy and related techniques like PALM, STORM and dSTORM use metastable dark states of a fluorophore for temporal separation of single molecules. https://www.leica-microsystems.com/science-lab/the-force-of-the-dark-side-embedding-media-for-gsdim-super-resolution-localization-microscopy/ Thu, 23 Jan 2014 18:49:00 +0000 Sebastian Bänfer https://www.leica-microsystems.com/10668 Laser Microdissection Video: How to Achieve Fast Precise Cutting Lines in Laser Microdissection This video explains the laser beam deflection principle of Leica Microsystems‘ laser microdissection systems (Leica LMD). The laser, the beam expander, motorized attenuator, motorized iris diaphragm for limiting the aperture for cutting and the scan unit are situated in the instrument’s special incident light axis. The scan unit consists of two prism wedges whose thickness and wedge angle are chosen so that the laser beam, after passing through the two prism wedges, is directed precisely through the center of the back focal plane of the objective. https://www.leica-microsystems.com/science-lab/video-how-to-achieve-fast-precise-cutting-lines-in-laser-microdissection/ Mon, 18 Nov 2013 14:32:00 +0000 M.Sc., Cornelia Gilbrich-Wille https://www.leica-microsystems.com/10717 EM Sample Preparation Rapid Immunohistochemical Diagnosis of Tobacco Mosaic Virus Disease by Microwave-assisted Plant Sample Preparation Immunoelectron microscopy is a powerful method to diagnose viral diseases and to study the distribution of the viral agent within plant cells and tissues. Nevertheless, current protocols for the immunological detection of viral diseases with transmission electron microscopy (TEM) in plants take between 3 and 6 days and are therefore not suited for rapid diagnosis of virus diseases in plants. In this study, we describe a method that allows rapid cytohistochemical detection of tobacco mosaic virus (TMV) in leaves of tobacco plants. https://www.leica-microsystems.com/science-lab/rapid-immunohistochemical-diagnosis-of-tobacco-mosaic-virus-disease-by-microwave-assisted-plant-sample-preparation/ Wed, 16 Oct 2013 20:36:00 +0000 Prof. Günther Zellnig, Stefan Möstl, Prof. Bernd Zechmann https://www.leica-microsystems.com/10912 EM Sample Preparation Brief Introduction to Contrasting for EM Sample Preparation Since the contrast in the electron microscope depends primarily on the differences in the electron density of the organic molecules in the cell, the efficiency of a stain is determined by the atomic weight of the stain attached to the biological structures. Consequently, the most widely used stains in electron microscopy are the heavy metals uranium and lead. https://www.leica-microsystems.com/science-lab/brief-introduction-to-contrasting-for-em-sample-preparation/ Wed, 02 Oct 2013 14:33:00 +0000 Dr. Ruwin Pandithage https://www.leica-microsystems.com/10224 EM Sample Preparation Brief Introduction to Glass Knifemaking for Electron and Light Microscope Applications Glass knives are used in an ultramicrotome to cut ultrathin slices of samples for electron and light microscope applications. For resin and for cryo sections (Tokuyasu samples) the knife edge must be extremely sharp, strong and stable. An important requirement for breaking glass knives of superior quality is the quality of the glass strips. Glass strips are produced from specially selected glass, the thickness and quality of which is precisely controlled. Only strict tolerances ensured by careful quality control allow breaking of two high quality knives from one square. https://www.leica-microsystems.com/science-lab/brief-introduction-to-glass-knifemaking-for-electron-and-light-microscope-applications/ Fri, 13 Sep 2013 15:49:00 +0000 Dr. Ruwin Pandithage https://www.leica-microsystems.com/10062 EM Sample Preparation Safe High Pressure Freezing of Infectious Microorganisms Especially in core EM-facilities one is confronted with material (microorganisms and cells) which are infectious. It is a must to follow the safety rules according to the National regulations. However even when safe laboratories are available it is very convenient to know that the applied high pressure freezing system is in itself safe. https://www.leica-microsystems.com/science-lab/safe-high-pressure-freezing-of-infectious-microorganisms/ Wed, 24 Jul 2013 14:10:00 +0000 PhD Dimitri Vanhecke, Dr. Benoit Zuber, Dr. Silvio D. Brugger, Daniel Studer https://www.leica-microsystems.com/9555 EM Sample Preparation Immersion Freezing of Suspended Particles and Cells for Cryo-Electron Microscopy Immersion freezing of thin aqueous specimens is an essential preparation technique for cryo-transmission electron microscopy (cryo-TEM), aiming to preserve fragile biological structures such as molecules and cells in their hydrated environment for a close-to-native visualization. For successful experiments, vitreous ice must be produced, surface contamination must be avoided, and, most important, the natural state of the structure must be preserved. https://www.leica-microsystems.com/science-lab/immersion-freezing-of-suspended-particles-and-cells-for-cryo-electron-microscopy/ Fri, 12 Apr 2013 11:48:00 +0000 Dr. Guenter Resch, Marlene Brandstetter, Lisa Königsmaier, Edith Urban, Angela M. Pickl-Herk https://www.leica-microsystems.com/7405 EM Sample Preparation Perusing Alternatives for Automated Staining of TEM Thin Sections Contrast in transmission electron microscopy (TEM) is mainly produced by electron scattering at the specimen: Structures that strongly scatter electrons are referred to as electron dense and appear as dark areas in the bright fi eld image, while structures which scatter fewer electrons appear bright (electron transparent). https://www.leica-microsystems.com/science-lab/perusing-alternatives-for-automated-staining-of-tem-thin-sections/ Mon, 04 Feb 2013 23:00:00 +0000 Nicole Fellner, Marlene Brandstetter, Karin Trimmel, Dr. Guenter Resch https://www.leica-microsystems.com/5734 EM Sample Preparation University of Wollongong Electron Microscopy Centre The University of Wollongong has a diverse range of materials research programs that includes metallurgy for mining, manufacturing, steel making and transport; polymers for solar cells, energy storage and bionic implants; and superconducting and electronic materials for commercialization, energy storage, telecommunications and medical applications. Electron microscopy is an integral part of this research, due to the chemical and structural information that can be provided down to the atomic scale. https://www.leica-microsystems.com/science-lab/university-of-wollongong-electron-microscopy-centre/ Thu, 17 Jan 2013 23:00:00 +0000 Darren Attard, Tony Romeo https://www.leica-microsystems.com/7972 EM Sample Preparation Brief Introduction to Critical Point Drying One of the uses of the Scanning Electron Microscope (SEM) is in the study of surface morphology in biological applications which requires the preservation of the surface details of a specimen. Samples for Electron Microscopy (EM) imaging need to be dried in order to be compatible with the vacuum in the microscope. The presence of water molecules will disturb the vacuum and with it the imaging. https://www.leica-microsystems.com/science-lab/brief-introduction-to-critical-point-drying/ Mon, 10 Dec 2012 23:00:00 +0000 Dr. Ruwin Pandithage https://www.leica-microsystems.com/5723 EM Sample Preparation Atomic Force Microscopy Study of a Stretched Impact Copolymer Atomic force microscopy (AFM) is a powerful tool in the suite of nanoscale characterization techniques that provides a variety of information including topography, mechanical properties, and electrical properties with nanoscale lateral and sub-nanometer vertical resolution. Cryoultramicrotomy is an essential tool for effective polymer sample preparation for atomic force microscopy (AFM) in order to get rid of the polymer skin from processing and to ensure a smooth surface for analysis. https://www.leica-microsystems.com/science-lab/atomic-force-microscopy-study-of-a-stretched-impact-copolymer/ Mon, 10 Dec 2012 23:00:00 +0000 Dr. Dalia G. Yablon, Jean Grabowski, Andy Haishung Tsou https://www.leica-microsystems.com/5738 EM Sample Preparation Dry Ultrathin Sectioning Combined With High Pressure Freezing We have used cultured UMR106-01 osteoblastic cells to investigate the process of bone mineralization. UMR106-01 cells as well as primary calvarial bone cells assembly spherical extracellular supramolecular protein-lipid complexes, termed biomineralization foci (BMF), in which the first crystals of hydroxyapatite mineral are deposited (Midura et al., 2004; Wang et al., 2004). A major difference between these culture models is the speed with which mineralization occurs, ranging from 12–16 days after plating for primary osteoblastic cells to 88 h for UMR106-01 cells. https://www.leica-microsystems.com/science-lab/dry-ultrathin-sectioning-combined-with-high-pressure-freezing/ Mon, 10 Dec 2012 23:00:00 +0000 Ph.D. Jeff P. Gorski, M.Sc. Nichole T. Huffman, Thérèse Hillman-Marti, Daniel Studer https://www.leica-microsystems.com/6742 Laser Microdissection Comparison of the RNA Quality of Native Tissue Samples with Tissue Samples after UV Laser Microdissection Due to its instability, RNA is generally more difficult to work with than DNA. RNA does not have the stability of the DNA double helix. To obtain the best results when dealing with RNA it is essential to start out with high-quality material and to carry out particularly careful quality control before and after processing. The so-called RIN (RNA Integrity Number) is an indicator of the quality of RNA. On a scale of 1–10, a RIN value of 1 indicates that the RNA is completely degraded and a RIN value of 10 that the RNA is fully intact. The higher the RIN number, the better the RNA quality. Wherever possible, material with high RIN numbers should always be used. https://www.leica-microsystems.com/science-lab/comparison-of-the-rna-quality-of-native-tissue-samples-with-tissue-samples-after-uv-laser-microdissection/ Sun, 30 Sep 2012 22:00:00 +0000 M.Sc., Cornelia Gilbrich-Wille https://www.leica-microsystems.com/4658 Super-Resolution Video Tutorial: How to Optimize Sample Preparation for GSD Microscopy This video tutorial presents a reliable way to produce samples for super-resolution GSD imaging with a special focus on the mounting step. Stable and flat mounting of the coverslip increases the performance of the overall system, leading to an improved resolution of the GSD image. https://www.leica-microsystems.com/science-lab/video-tutorial-how-to-optimize-sample-preparation-for-gsd-microscopy/ Fri, 13 Jan 2012 10:09:00 +0000 Beate Braun, Dr. Marko Lampe https://www.leica-microsystems.com/10998 EM Sample Preparation Microwave Assisted Rapid Diagnosis of Plant Virus Diseases by Transmission Electron Microscopy Investigations of ultrastructural changes induced by viruses are often necessary to clearly identify viral diseases in plants. With conventional sample preparation for transmission electron microscopy (TEM) such investigations can take several days and are therefore not suited for a rapid diagnosis of plant virus diseases. Microwave fixation can be used to drastically reduce sample preparation time for TEM investigations with similar ultrastructural results as observed after conventionally sample preparation https://www.leica-microsystems.com/science-lab/microwave-assisted-rapid-diagnosis-of-plant-virus-diseases-by-transmission-electron-microscopy/ Fri, 14 Oct 2011 03:28:00 +0000 Prof. Bernd Zechmann, Prof. Günther Zellnig https://www.leica-microsystems.com/11027 EM Sample Preparation Neuroscience Focussed Ion Beam Milling and Scanning Electron Microscopy of Brain Tissue This protocol describes how biological samples, like brain tissue, can be imaged in three dimensions using the focussed ion beam/scanning electron microscope (FIB/SEM). The samples are fixed with aldehydes, heavy metal stained using osmium tetroxide and uranyl acetate. They are then dehydrated with alcohol and infiltrated with resin, which is then hardened. https://www.leica-microsystems.com/science-lab/focussed-ion-beam-milling-and-scanning-electron-microscopy-of-brain-tissue/ Wed, 06 Jul 2011 16:17:00 +0000 https://www.leica-microsystems.com/10831 CLEM EM Sample Preparation Advanced Correlative Light/Electron Microscopy: Current Methods and New Developments Using Tokuyasu Cryosections Microscopy is an essential tool for analysis of cellular structures and function. With the advent of new fluorescent probes and super-resolution light microscopy techniques, the study of dynamic processes in living cells has been greatly facilitated. Fluorescence light microscopy provides analytical, quantitative, and three-dimensional (3D) data with emphasis on analysis of live cells using fluorescent markers. https://www.leica-microsystems.com/science-lab/advanced-correlative-lightelectron-microscopy-current-methods-and-new-developments-using-tokuyasu-cryosections/ Sun, 20 Dec 2009 09:11:00 +0000 https://www.leica-microsystems.com/7970 CLEM EM Sample Preparation CLEM: Combining the Strengths of Light and Electron Microscopy In recent years light microscopy studies have been dominated by live cell imaging while electron microscopy has been used for high-resolution studies. Latterly, there has been increasing interest in combining these techniques. This combination is called Correlative Light Electron Microscopy (CLEM). Due to the high resolution made possible by electron microscopy, artefacts induced during preparation of a sample can, however, also be clearly seen. https://www.leica-microsystems.com/science-lab/clem-combining-the-strengths-of-light-and-electron-microscopy/ Mon, 09 Apr 2007 22:00:00 +0000 Professor Paul Verkade