Collection of Continuous Rotation MicroED Data from Ion Beam-Milled Crystals of Any Size

Structure. 2019 Mar 5;27(3):545-548.e2. doi: 10.1016/j.str.2018.12.003. Epub 2019 Jan 17.

Abstract

Microcrystal electron diffraction (MicroED) allows for macromolecular structure solution from nanocrystals. To create crystals of suitable size for MicroED data collection, sample preparation typically involves sonication or pipetting a slurry of crystals from a crystallization drop. The resultant crystal fragments are fragile and the quality of the data that can be obtained from them is sensitive to subsequent sample preparation for cryoelectron microscopy as interactions in the water-air interface can damage crystals during blotting. Here, we demonstrate the use of a focused ion beam to generate lamellae of macromolecular protein crystals for continuous rotation MicroED that are of ideal thickness, easy to locate, and require no blotting optimization. In this manner, crystals of nearly any size may be scooped and milled to desired dimensions prior to data collection, thus streamlining the methodology for sample preparation for MicroED.

Keywords: Electron cryo-microscopy (cryoEM); FIB-SEM; MicroED; electron crystallography; electron diffraction.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cryoelectron Microscopy
  • Crystallography
  • Macromolecular Substances / chemistry*
  • Models, Molecular
  • Particle Size

Substances

  • Macromolecular Substances