相关实验视频

Updated: Sep 8, 2025

Preparing Lamellae from Vitreous Biological Samples Using a Dual-Beam Scanning Electron Microscope for Cryo-Electron Tomography

Preparing Lamellae from Vitreous Biological Samples Using a Dual-Beam Scanning Electron Microscope for Cryo-Electron Tomography

Published on: August 5, 2021

在CEMOVIS的高分辨率的in situ冷EM重建

Johannes Elferich¹, Marek Kaminek², Lingli Kong³

¹Howard Hughes Medical Institute, University of Massachusetts Chan Medical School, Worcester, MA, USA.

|June 24, 2025

概括

玻璃体截面的冷电子显微镜 (CEMOVIS) 提供了一个高分辨率的成像替代方案,而不是聚焦离子束 (FIB) 削. 这种方法提供了显著更大的可图像面积,在很大程度上保持完整的分子结构,与FIB磨砂样品相比.

科学领域:

结构生物学结构生物学
细胞生物学细胞生物学
显微镜技术显微镜技术

背景情况:

使用冷电子显微镜 (cryo-EM) 进行细胞和组织的高分辨率成像,需要电子透明样本 (厚度≤500nm).
聚焦离子束 (FIB) 研磨是制备冷电磁样品 (lamellae) 的标准,但它涉及材料损失,有限的可图像面积和离子束损伤.
玻璃体截面的冷电子显微镜 (CEMOVIS) 为样品制备提供了FIB削的潜在替代方案.

研究的目的:

评估CEMOVIS作为FIB削的替代品,用于冷-EM样本制备.
评估CEMOVIS提供的质量和可图像区域.
为了证明从CEMOVIS样本中获得高分辨率结构信息的可行性.

主要方法:

使用玻璃体截面冷电子显微镜 (CEMOVIS) 制备样本.
对玻璃体截面的分析,承认压缩,剪切和裂等潜在的损伤.
应用二维模板匹配用于结构重建.

主要成果:

通过CEMOVIS,可以将样品连续切割,与FIB粉碎的片相比,可以获得显著更多的可图像面积.
尽管切割引起的损伤,玻璃体部分内的分子结构在许多地区基本完好无损.
从Saccharomyces cerevisiae细胞中重建60S核糖体子单元的近原子分辨率是使用CEMOVIS数据实现的.

结论:

塞莫维斯是一种可行的FIB削替代品,用于冷-EM样本制备,在可图像面积方面具有明显的优势.
该技术保持了分子完整性,足以进行高分辨率的结构研究.
塞莫维斯扩大了细胞结构的冷电磁分析的可能性,特别是在大型成像研究中.

关键词:

冷电子显微镜的使用方法电子断层扫描是一种电子断层扫描.影像成像技术影像成像技术整合性结构生物学整合性结构生物学结构的确定结构的确定

更多相关视频

A Robust Single-Particle Cryo-Electron Microscopy cryo-EM Processing Workflow with cryoSPARC, RELION, and Scipion

A Robust Single-Particle Cryo-Electron Microscopy cryo-EM Processing Workflow with cryoSPARC, RELION, and Scipion

Published on: January 31, 2022

Visualization of Organelles In Situ by Cryo-STEM Tomography

Visualization of Organelles In Situ by Cryo-STEM Tomography

Published on: June 23, 2023

相关概念视频

Cryo-electron Microscopy

Cryo-electron Microscopy

Conventional electron microscopy (EM) involves dehydration, fixation, and staining of biological samples, which distorts the native state of biological molecules and results in several artifacts. Also, the high-energy electron beam damages the sample and makes it difficult to obtain high-resolution images. These issues can be addressed using cryo-EM, which uses frozen samples and gentler electron beams. The technique was developed by Jacques Dubochet, Joachim Frank, and Richard Henderson, for...

Electron Microscope Tomography and Single-particle Reconstruction

Electron Microscope Tomography and Single-particle Reconstruction

Transmission electron microscopy (TEM) can be used to determine the 3D structure of biological samples with the help of techniques such as electron microscope tomography and single-particle reconstruction. While single-particle reconstruction can examine macromolecules and macromolecular complexes in vitro conditions only, tomography permits the study of cell components or small cells in vivo.
Electron Tomography
Electron tomography can be performed either in TEM or STEM (scanning transmission...