title: Deformed grids for single-particle cryo-electron microscopy of specimens exhibiting a preferred orientation.
abstract: For biological samples showing a preferred orientation on the carbon support film of an electron microscope (EM) grid, accurate three-dimensional (3D) reconstructions by single-particle cryo-EM require data collection in which the specimen grids are tilted in the microscope, to obtain adequate numbers of particles that cover the high-degree angular distribution. However, image drift caused by the electron beam interacting with the cryo specimen becomes severe when grids are tilted to high angles (>30°). We produced deformed grids by applying a deliberate mechanical deformation to EM grids containing a thin carbon film supported by a thick holey carbon film. We applied cryo-EM using deformed grids to the isolated cardiac ryanodine receptor, an ion channel complex known to assume a preferred orientation on the carbon support film. These grids contained more particles having high Euler angle orientations without the need to tilt the specimen grids. Meanwhile, the drifting that was apparent in the images was reduced from that typical of images from tilted regular EM grids. This was achieved by imaging particles in holes close to the deformed areas, where carbon films were locally bent, offering planes of inclination with various angles. The deformed grids improve the efficiency and quality of data collection for single-particle cryo-EM of samples showing a limited range of orientations.
authors.affiliation: Wadsworth Center, New York State Department of Health, Albany, NY 12201, USA.
authors.forename: Ying
authors.initials: Y
authors.lastname: Liu
authors.forename: Xing
authors.initials: X
authors.lastname: Meng
authors.forename: Zheng
authors.initials: Z
authors.lastname: Liu
mesh_terms: D002244:Carbon / Q000737:chemistry*; D020285:Cryoelectron Microscopy; D007091:Image Processing, Computer-Assisted; D021621:Imaging, Three-Dimensional / Q000379:methods*; D013048:Specimen Handling
chemical_list: D002244:Carbon
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