title: An improved cryo-FIB method for fabrication of frozen hydrated lamella.
abstract: Cryo-electron tomography (cryo-ET) provides great insights into the ultrastructure of cells and tissues in their native state and provides a promising way to study the in situ 3D structures of macromolecular complexes. However, this technique has been limited on the very thin specimen, which is not applicable for most cells and tissues. Besides cryo-sectioning approach, cryo focused ion beam (cryo-FIB) appeared recently to achieve 'artifact-free' thin frozen hydrated lamella via fabrication. Considering that the current cryo-FIB methods need modified holders or cartridges, here, with a "D-shaped" molybdenum grid and a specific shutter system, we developed a simple cryo-FIB approach for thin frozen hydrated lamella fabrication, which fits both standard transmission cryo-electron microscopes with side-entry cryo-holders and state-of-the-art ones with AutoGrids. Our approach will expand the usage of cryo-FIB approach in many labs.
authors.affiliation: Center for Biological Imaging, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Chaoyang District, Beijing 100101, China.
authors.forename: Jianguo
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Center for Biological Imaging, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Chaoyang District, Beijing 100101, China.
authors.forename: Gang
authors.initials: G
authors.lastname: Ji
authors.affiliation: Center for Biological Imaging, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Chaoyang District, Beijing 100101, China.
authors.forename: Xiaojun
authors.initials: X
authors.lastname: Huang
authors.affiliation: Center for Biological Imaging, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Chaoyang District, Beijing 100101, China. Electronic address: emlb@ibp.ac.cn.
authors.forename: Wei
authors.initials: W
authors.lastname: Xu
authors.affiliation: Center for Biological Imaging, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Chaoyang District, Beijing 100101, China; National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Chaoyang District, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing, China. Electronic address: feisun@sun5.ibp.ac.cn.
authors.forename: Fei
authors.initials: F
authors.lastname: Sun
mesh_terms: D000818:Animals; D017165:Ascaris suum / Q000166:cytology; D020285:Cryoelectron Microscopy / Q000295:instrumentation / Q000379:methods*; D055032:Electron Microscope Tomography / Q000295:instrumentation / Q000379:methods*; D004926:Escherichia coli / Q000648:ultrastructure; D005347:Fibroblasts / Q000648:ultrastructure; D005629:Frozen Sections / Q000379:methods*; D008297:Male; D051379:Mice; D061987:Sf9 Cells / Q000648:ultrastructure; D013094:Spermatozoa / Q000648:ultrastructure; D018411:Spodoptera / Q000166:cytology
keywords: Cell ultrastructure; Cryo-electron tomography; Cryo-focused ion beam; Frozen hydrated lamella; Molybdenum grid