Measuring the optimal exposure for single particle cryo-EM using a 2.6 Å reconstruction of rotavirus VP6

By Tim Grant, N. Grigorieff. eLife, 2015

814 citations recorded by the source corpus.

Biological specimens suffer radiation damage when imaged in an electron microscope, ultimately limiting the attainable resolution. At a given resolution, an optimal exposure can be defined that maximizes the signal-to-noise ratio in the image. Using a 2.6 Å resolution single particle cryo-EM reconstruction of rotavirus VP6, determined from movies recorded with a total exposure of 100 electrons/Å2, we obtained accurate measurements of optimal exposure values over a wide range of resolutions. At low and intermediate resolutions, our measured values are considerably higher than obtained previously for crystalline specimens, indicating that both images and movies should be collected with higher exposures than are generally used. We demonstrate a method of using our optimal exposure values to filter movie frames, yielding images with improved contrast that lead to higher resolution reconstructions. This ‘high-exposure’ technique should benefit cryo-EM work on all types of samples, especially those of relatively low-molecular mass. DOI: http://dx.doi.org/10.7554/eLife.06980.001

Subfields: Genetics, Infectious Diseases, Structural Biology

View Paper: https://www.semanticscholar.org/paper/2ab09e0bd6be8101cc84205102fbd14615e9de20

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Citations in the corpus

AreTomo: An integrated software package for automated marker-free, motion-corrected cryo-electron tomographic alignment and reconstruction (2022, methodology): “(Grant and Grigorieff, 2015) for cryoET by treating a tilt series as if it were a single movie containing all tilt images sorted in the order of acquisition.”
Visualization of two architectures in class-II CAP-dependent transcription activation (2019, methodology): “physical drift were corrected followed by dose-weighing using the Unblur algorithm [33].”
A Bayesian approach to beam-induced motion correction in cryo-EM single-particle analysis (2018, methodology): “This assumption is further supported by the fact that β-galactosidase is also the only dataset on which traditional polishing applied after Unblur produces a better reconstruction than running MotionCor2 alone.”
Accelerated cryo-EM structure determination with parallelisation using GPUs in RELION-2 (2016, methodology): “Super-resolution 8 k (cid:2) 8 k micrograph movies with 38 frames were submitted to initial beam-induced motion correction using UNBLUR ( Grant and Grigorieff, 2015 ).”