A second generation apparatus for time-resolved electron cryo-microscopy using stepper motors and electrospray
A second generation apparatus for time-resolved electron cryo-microscopy using stepper motors and electrospray
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DOI：
10.1016/j.jsb.2003.09.027
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发表时间：
2003-10-01
影响因子：
3
通讯作者：
Trinick, J
中科院分区：
文献类型：
--
作者：
White, HD;Thirumurugan, K;Trinick, J
我们在这里描述了用电子冷冻显微镜研究大分子及其复合物的瞬态反应构象的第二代装置。反应在开始后几毫秒到几十秒的时间内被快速冻结。利用计算机控制的微型步进电机对电子显微镜栅格进行印迹并将其冷冻在液态乙烷中。为了获得最快的时间分辨率，在一个含有一种反应物薄膜的网格中，在冷冻前喷洒含有第二种反应物的小液滴。喷雾是用电产生的（电喷雾），从每个网格所需的1-2摩尔溶液中产生直径小于1微米的密集液滴云。第二种方法是先用湍流混合两种溶液，然后在冻结前进行吸渍，用于时间过程为bbb51 s的反应。(C) 2003 Elsevier Inc.版权所有。
We describe here a second generation apparatus for studying transient reaction conformations in macromolecules and their complexes by electron cryo-microscopy. Reactions are trapped by rapid freezing in times ranging from a few milliseconds to tens of seconds after initiation. Blotting of the electron microscope grid and freezing it in liquid ethane uses computer controlled micro-stepping motors. For the fastest time resolution, a blotted grid containing a thin film of one reactant is sprayed with small droplets containing a second reactant just before freezing. The spray is produced electrically (electrospray), which gives a dense cloud of droplets < 1 mum in diameter from the 1-2 mul of solution required per grid. A second method in which two solutions are first mixed by turbulent flow and then blotted prior to freezing is used for reactions with time courses > 1 s. (C) 2003 Elsevier Inc. All rights reserved.