Automatic estimation and correction of anisotropic magnification distortion in electron microscopes.
We demonstrate a significant anisotropic magnification distortion, found on an FEI Titan Krios microscope and affecting magnifications commonly used for data acquisition on a Gatan K2 Summit detector. We describe a program (mag_distortion_estimate) to automatically estimate anisotropic magnification distortion from a set of images of a standard gold shadowed diffraction grating. We also describe a program (mag_distortion_correct) to correct for the estimated distortion in collected images. We demonstrate that the distortion present on the Titan Krios microscope limits the resolution of a set of rotavirus VP6 images to ∼7 Å, which increases to ∼3 Å following estimation and correction of the distortion. We also use a 70S ribosome sample to demonstrate that in addition to affecting resolution, magnification distortion can also interfere with the classification of heterogeneous data.
Janelia Research Campus, Howard Hughes Medical Institute, 19700 Helix Drive, Ashburn, VA 20147, USA.
Timothy
T
Grant
Janelia Research Campus, Howard Hughes Medical Institute, 19700 Helix Drive, Ashburn, VA 20147, USA. Electronic address: niko@grigorieff.org.
Nikolaus
N
Grigorieff
D016880:Anisotropy; D020285:Cryoelectron Microscopy / Q000295:instrumentation / Q000379:methods*; D006046:Gold / Q000032:analysis*; D007089:Image Enhancement / Q000379:methods*
D006046:Gold
Electron cryo-microscopy; Image correction; Magnification anisotropy; Resolution