相关实验视频
Updated: Jan 14, 2026

09:04
Miniaturized Sample Preparation for Transmission Electron Microscopy
Published on: July 27, 2018
20.5K
在冷EMEM中,超越蛋白质扩散到空气-水接口
Anastasiia Gusach1, Kasim Sader2, Christopher J Russo1
1Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 0QH, United Kingdom.
概括
研究人员开发了一种用于电子冷显微镜 (cryoEM) 的快速样本准备技术. 这种方法快速玻璃化样品,防止蛋白质扩散到空气-水界面,并保留原子结构.
科学领域:
- 结构生物学 结构生物学
- 生物物理学的生物物理.
- 材料科学 材料科学 材料科学
背景情况:
- 空气-水接口是冷EM样本制备中的一个主要挑战.
- 蛋白质扩散到这个接口可以导致变性和文物.
- 玻璃化对于保持生物分子结构在其原始状态至关重要.
研究的目的:
- 开发一种用于冷EM中快速样本玻璃化的新方法.
- 为了克服空气-水接口所带来的局限性.
- 为了在样本准备过程中保留蛋白质的原生原子结构.
主要方法:
- 在高速度 (几百米/秒) 喷picoliter滴到一个液体乙涂层,预冷的支持.
- 同时滴滴崩和玻璃化进入无形阶段在微秒内.
- 使用快速结技术绕过空气-水接口相互作用.
主要成果:
- 实现了比空气-水接口的蛋白质扩散率更快的玻璃化.
- 在玻璃化样本中保存了蛋白质的原子结构.
- 取消了标本对接口的粘附,通过断层图像重建证实了这一点.
- 标识了样品厚度和粒子方向作为新的限制因素.
结论:
- 证明了快速玻璃化的物理可能性,以减轻空气-水接口效应.
- 这种技术为开发新的冷EM样本准备仪器提供了基础.
- 未来的工作应该集中在控制样品厚度和粒子方向,以改善冷EM数据.
相关概念视频
Protein Diffusion in the Membrane
5.4K
Proteins show rotational as well as lateral diffusion across the membrane. The lateral diffusion of proteins was confirmed through the cell fusion experiment where mouse and human cells were fused, resulting in hybrid cells. When the human and mouse cells fused, the specific membrane proteins on human and mouse cells were marked with the red and green-fluorescent markers, respectively. Initially, the red and green fluorescence was located on the respective hemisphere of the cell. As time...
5.4K
Cryo-electron Microscopy
4.1K
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...
4.1K
Protein Dynamics in Living Cells
2.6K
Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...
2.6K

