Microscopic charge fluctuations cause minimal contrast loss in cryoEM.
Microscopic charge fluctuations cause minimal contrast loss in cryoEM.
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DOI：
10.1016/j.ultramic.2018.01.011
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发表时间：
2018-04
期刊：
影响因子：
2.2
通讯作者：
Henderson R
中科院分区：
文献类型：
--
作者：
Russo CJ;Henderson R
提出了低剂量冷冻电镜中电荷波动的物理解释。我们量化了冷冻电镜样本上的蜂群充电现象。我们测量由电荷波动引起的包络函数。这些波动的影响在冷冻电镜中可以忽略不计。在高度散焦的透射电子显微照片中看到的波动粒度或“蜂群”效应是由照明光束在样品中产生的微观电荷波动引起的。在高分辨率单粒子电子冷冻显微镜 (cryoEM) 领域，人们担心这种波动的电荷可能会导致散焦相关的托恩环衰落，从而降低最终图像的质量。在本文中，我们分析了嵌入非晶冰中的金纳米颗粒图像中 (111) 反射的 2.35 Å 条纹。我们表明，与纯金箔相比，晶格条纹存在少量但可检测的散焦相关模糊。与绝缘冷冻水样本上的波动电荷相关的横向电场使电子束局部偏转并导致图像模糊。扰动很小，使 10μm 散焦处 2.35 Å 反射的振幅降低约 7%（强度降低 14%）。对于 2–4μm 范围内的较小散焦值以及冷冻电镜中常见的分辨率，假设使用场发射枪 (FEG) 进行照明，则对于所有合理的冷冻电镜成像条件，源不相干和蜂群效应的影响可以忽略不计。这使得由于辐射损伤而导致的样本物理移动成为突出问题，也是冷冻电镜显微照片中对比度损失的主要来源。
A physical account of charge fluctuations in low-dose cryoEM is presented. We quantify the bee swarm charging phenomenon on cryoEM specimen. We measure the envelope function caused by charge fluctuations. The effects of these fluctuations are negligible in cryoEM. The fluctuating granularity or “bee swarm” effect seen in highly defocussed transmission electron micrographs is caused by microscopic charge fluctuations in the specimen created by the illuminating beam. In the field of high-resolution single particle electron cryomicroscopy (cryoEM), there has been a concern that this fluctuating charge might cause defocus-dependent Thon ring fading which would degrade the final image. In this paper, we have analysed the 2.35 Å fringes from the (111) reflection in images of gold nanoparticles embedded in amorphous ice. We show that there is a small, yet detectable amount of defocus-dependent blurring of the lattice fringes when compared with those from a pure gold foil. The transverse electric field associated with the fluctuating charges on the insulating frozen water specimen deflects the electron beam locally and causes image blurring. The perturbation is small, decreasing the amplitude of the 2.35 Å reflection at 10 µm defocus by about 7% (intensity by 14%). For smaller defocus values in the range 2–4 µm and for resolutions that are typical in cryoEM, the effects of source incoherence and the bee swarm effect are negligible for all reasonable cryoEM imaging conditions, assuming that a field emission gun (FEG) is used for illumination. This leaves physical movement of the specimen due to radiation damage as the outstanding problem and the major source of contrast loss in cryoEM micrographs.
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影响因子：
2.8
作者：
Glaeser, RM;Downing, KH
通讯作者：
Downing, KH
影响因子：
2.2
作者：
McMullan, G.;Vinothkumar, K. R.;Henderson, R.
通讯作者：
Henderson, R.
DOI：
10.1073/pnas.1418377111
发表时间：
2014-11-04
影响因子：
11.1
作者：
Danev, Radostin;Buijsse, Bart;Baumeister, Wolfgang
通讯作者：
Baumeister, Wolfgang
影响因子：
2.8
作者：
Downing, KH;McCartney, MR;Glaeser, RM
通讯作者：
Glaeser, RM
影响因子：
16.6
作者：
Khoshouei M;Radjainia M;Baumeister W;Danev R
通讯作者：
Danev R