Graphene Oxide‐Supported Microwell Grids for Preparing Cryo‐EM Samples with Controlled Ice Thickness

Abstract

Cryogenic‐electron microscopy (cryo‐EM) is the preferred method to determine 3D structures of proteins and to study diverse material systems that intrinsically have radiation or air sensitivity. Current cryo‐EM sample preparation methods provide limited control over the sample quality, which limits the efficiency and high throughput of 3D structure analysis. This is partly because it is difficult to control the thickness of the vitreous ice that embeds specimens, in the range of nanoscale, depending on the size and type of materials of interest. Thus, there is a need for fine regulation of the thickness of vitreous ice to deliver consistent high signal‐to‐noise ratios for low‐contrast biological specimens. Herein, an advanced silicon‐chip‐based device is developed which has a regular array of micropatterned holes with a graphene oxide (GO) window on freestanding silicon nitride (SixNy). Accurately regulated depths of micropatterned holes enable precise control of vitreous ice thickness. Furthermore, GO window with affinity for biomolecules can facilitate concentration of the sample molecules at a higher level. Incorporation of micropatterned chips with a GO window enhances cryo‐EM imaging for various nanoscale biological samples including human immunodeficiency viral particles, groEL tetradecamers, apoferritin octahedral, aldolase homotetramer complexes, and tau filaments, as well as inorganic materials.

Facile preparation of graphene–graphene oxide liquid cells and their application in liquid-phase STEM imaging of Pt atoms

“…The resolution of LP-STEM is better than that of LP-TEM, and aberrationcorrected STEM enables imaging of the atomic structures of samples and even single atoms in liquid, whereas scanning speed limits the temporal resolution. 2,17,26) Generally, the basal plane of GO produced by chemical oxidation methods, like the modified Hummers method, predominantly contains epoxy and hydroxyl functional groups, [27][28][29][30][31][32] resulting in strong water adhesion (i.e., hydrophilicity). This is expected to increase the production yield of liquid sample pockets on GO relative to graphene.…”