Department of Biomedical-Chemical Engineering, The Catholic University of Korea; Department of Biotechnology, The Catholic University of Korea.
School of Chemical and Biological Engineering, and Institute of Chemical Processes, Seoul National University; Center for Nanoparticle Research, Institute of Basic Science (IBS).
J Vis Exp. 2022 Apr 21(182). doi: 10.3791/63739.
A major limitation for the efficient and high-throughput structure analysis of biomolecules using cryogenic electron microscopy (cryo-EM) is the difficulty of preparing cryo-EM samples with controlled ice thickness at the nanoscale. The silicon (Si)-based chip, which has a regular array of micro-holes with graphene oxide (GO) window patterned on a thickness-controlled silicon nitride (SixNy) film, has been developed by applying microelectromechanical system (MEMS) techniques. UV photolithography, chemical vapor deposition, wet and dry etching of the thin film, and drop-casting of 2D nanosheet materials were used for mass-production of the micro-patterned chips with GO windows. The depth of the micro-holes is regulated to control the ice thickness on-demand, depending on the size of the specimen for cryo-EM analysis. The favorable affinity of GO toward biomolecules concentrates the biomolecules of interest within the micro-hole during cryo-EM sample preparation. The micro-patterned chip with GO windows enables high-throughput cryo-EM imaging of various biological molecules, as well as inorganic nanomaterials.
使用低温电子显微镜（cryo-EM）对生物分子进行高效、高通量结构分析的一个主要限制是，很难在纳米尺度上制备具有受控冰厚的 cryo-EM 样品。基于硅（Si）的芯片采用微电子机械系统（MEMS）技术开发，在厚度可控的氮化硅（SixNy）薄膜上具有图案化有氧化石墨烯（GO）窗口的规则微孔阵列。该芯片通过使用紫外光刻、薄膜化学气相沉积、薄膜的湿法和干法刻蚀以及二维纳米片材料的滴铸，实现了带有 GO 窗口的微图案化芯片的大规模生产。通过调节微孔的深度，可以根据 cryo-EM 分析用的样品尺寸来按需控制冰的厚度。GO 对生物分子的有利亲和力可在 cryo-EM 样品制备过程中将感兴趣的生物分子浓缩在微孔内。带有 GO 窗口的微图案化芯片可实现各种生物分子以及无机纳米材料的高通量 cryo-EM 成像。
