Comparison of different preparation methods of biological samples for FIB milling and SEM investigation. 2009

V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
Department of Biology, University of Ljubljana, Vecna pot 111, SI-1000 Ljubljana, Slovenia.

When a new approach in microscopy is introduced, broad interest is attracted only when the sample preparation procedure is elaborated and the results compared with the outcome of the existing methods. In the work presented here we tested different preparation procedures for focused ion beam (FIB) milling and scanning electron microscopy (SEM) of biological samples. The digestive gland epithelium of a terrestrial crustacean was prepared in a parallel for FIB/SEM and transmission electron microscope (TEM). All samples were aldehyde-fixed but followed by different further preparation steps. The results demonstrate that the FIB/SEM samples prepared for conventional scanning electron microscopy (dried) is suited for characterization of those intracellular morphological features, which have membranous/lamellar appearance and structures with composition of different density as the rest of the cell. The FIB/SEM of dried samples did not allow unambiguous recognition of cellular organelles. However, cellular organelles can be recognized by FIB/SEM when samples are embedded in plastic as for TEM and imaged by backscattered electrons. The best results in terms of topographical contrast on FIB milled dried samples were obtained when samples were aldehyde-fixed and conductively stained with the OTOTO method (osmium tetroxide/thiocarbohydrazide/osmium tetroxide/thiocarbohydrazide/osmium tetroxide). In the work presented here we provide evidence that FIB/SEM enables both, detailed recognition of cell ultrastructure, when samples are plastic embedded as for TEM or investigation of sample surface morphology and subcellular composition, when samples are dried as for conventional SEM.

UI MeSH Term Description Entries
D008855 Microscopy, Electron, Scanning Microscopy in which the object is examined directly by an electron beam scanning the specimen point-by-point. The image is constructed by detecting the products of specimen interactions that are projected above the plane of the sample, such as backscattered electrons. Although SCANNING TRANSMISSION ELECTRON MICROSCOPY also scans the specimen point by point with the electron beam, the image is constructed by detecting the electrons, or their interaction products that are transmitted through the sample plane, so that is a form of TRANSMISSION ELECTRON MICROSCOPY. Scanning Electron Microscopy,Electron Scanning Microscopy,Electron Microscopies, Scanning,Electron Microscopy, Scanning,Electron Scanning Microscopies,Microscopies, Electron Scanning,Microscopies, Scanning Electron,Microscopy, Electron Scanning,Microscopy, Scanning Electron,Scanning Electron Microscopies,Scanning Microscopies, Electron,Scanning Microscopy, Electron
D009942 Organometallic Compounds A class of compounds of the type R-M, where a C atom is joined directly to any other element except H, C, N, O, F, Cl, Br, I, or At. (Grant & Hackh's Chemical Dictionary, 5th ed) Metallo-Organic Compound,Metallo-Organic Compounds,Metalloorganic Compound,Organometallic Compound,Metalloorganic Compounds,Compound, Metallo-Organic,Compound, Metalloorganic,Compound, Organometallic,Compounds, Metallo-Organic,Compounds, Metalloorganic,Compounds, Organometallic,Metallo Organic Compound,Metallo Organic Compounds
D004064 Digestive System A group of organs stretching from the MOUTH to the ANUS, serving to breakdown foods, assimilate nutrients, and eliminate waste. In humans, the digestive system includes the GASTROINTESTINAL TRACT and the accessory glands (LIVER; BILIARY TRACT; PANCREAS). Ailmentary System,Alimentary System
D004848 Epithelium The layers of EPITHELIAL CELLS which cover the inner and outer surfaces of the cutaneous, mucus, and serous tissues and glands of the body. Mesothelium,Epithelial Tissue,Mesothelial Tissue,Epithelial Tissues,Mesothelial Tissues,Tissue, Epithelial,Tissue, Mesothelial,Tissues, Epithelial,Tissues, Mesothelial
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D013194 Staining and Labeling The marking of biological material with a dye or other reagent for the purpose of identifying and quantitating components of tissues, cells or their extracts. Histological Labeling,Staining,Histological Labelings,Labeling and Staining,Labeling, Histological,Labelings, Histological,Stainings
D016707 Tissue Fixation The technique of using FIXATIVES in the preparation of cytologic, histologic, or pathologic specimens for the purpose of maintaining the existing form and structure of all the constituent elements. Fixation, Tissue
D033321 Isopoda One of the largest orders of mostly marine CRUSTACEA, containing over 10,000 species. Like AMPHIPODA, the other large order in the superorder Peracarida, members are shrimp-like in appearance, have sessile compound eyes, and no carapace. But unlike Amphipoda, they possess abdominal pleopods (modified as gills) and their bodies are dorsoventrally flattened. Isopods,Isopod,Isopodas

Related Publications

V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
January 2010, The Journal of clinical dentistry,
V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
July 2023, Microscopy and microanalysis : the official journal of Microscopy Society of America, Microbeam Analysis Society, Microscopical Society of Canada,
V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
January 2023, Methods in cell biology,
V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
September 2022, Microscopy and microanalysis : the official journal of Microscopy Society of America, Microbeam Analysis Society, Microscopical Society of Canada,
V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
January 2004, Micron (Oxford, England : 1993),
V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
October 2018, Faraday discussions,
V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
January 2014, Methods in molecular biology (Clifton, N.J.),
V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
October 2020, Journal of imaging,
V Leser, and D Drobne, and Z Pipan, and M Milani, and F Tatti
January 2019, Methods in cell biology,
Copied contents to your clipboard!