![]() Hair dryer (e.g., Steinel HG 2000 from VWR International) or Leica EM CTD drying platform (available from mid-2011 on) ![]() This is stronger and renders the films more hydrophilic, but should only be used for very stable films such as Quantifoil. Instrument built according to Aebi and Pollard (1987) This provides more gentle discharge it is used for thin continuous carbon. 2007) or purchased from EM suppliers (e.g., Quantifoil Micro Tools GmbH, Jena, Germany, Glow discharge unit, either:īal-Tec SCD005 sputter coater (now Leica Microsystems, Vienna, Austria) with the Au target removed ![]() Made with a high vacuum evaporator ( Grassucci et al. Both continuous as well as perforated (holey) carbon films can be (2010) for an in-depth discussion of the best choice of grids. See Table 1 for examples and Dobro et al. Troubleshooting issues concerning the operation of the GP in particular, as well as common problems in immersionįreezing encountered on manual and semiautomatic instruments, are addressed.Ĭryo-grid boxes with lid, either 15- × 15-mm square (e.g., Ted Pella 160-44) or 14-mm diameter round (e.g., Leica MicrosystemsĮach of these boxes can accommodate four grids.ĮM grids with mesh size and support film suitable for your experiment It also provides details on how to make the most of the special features of this instrument to obtain good specimens and reproducible Of issues of general importance for cryo-EM, such as the properties of the sample and the pretreatment of the specimen carrier. Organelles, or small cells suspended in an aqueous solution, using the new Leica “EM GP” grid plunger. ![]() Of biological samples, such as purified protein complexes, viruses, liposomes, synthetic cytoskeletal filaments, isolated This protocol describes immersion freezing For successful experiments, vitreous ice must be produced, surface contamination must beĪvoided, and, most important, the natural state of the structure must be preserved. (cryo-TEM), aiming to preserve fragile biological structures such as molecules and cells in their hydrated environment forĪ close-to-native visualization. Immersion freezing of thin aqueous specimens is an essential preparation technique for cryo-transmission electron microscopy ![]()
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