A photoprotection strategy for microsecond-resolution single-molecule fluorescence spectroscopy
A photoprotection strategy for microsecond-resolution single-molecule fluorescence spectroscopy
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DOI：
10.1038/nmeth.1553
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发表时间：
2011-02-01
期刊：
影响因子：
48
通讯作者：
Munoz, Victor
中科院分区：
文献类型：
--
作者：
Campos, Luis A.;Liu, Jianwei;Munoz, Victor
由于染料闪烁和漂白，当前单分子荧光技术的时间分辨率限于毫秒。在这里，我们介绍了一种光保护策略，提供微秒分辨率相结合的有效的三重淬灭氧和Trolox通过氧自由基清除剂半胱胺漂白最小化。使用这种方法，我们解决了两个蛋白质的单分子微秒构象波动：两个国家的文件夹α-血影蛋白SH 3结构域和超快下坡文件夹BBL。
Time resolution of current single-molecule fluorescence techniques is limited to milliseconds because of dye blinking and bleaching. Here we introduce a photoprotection strategy that affords microsecond resolution by combining efficient triplet quenching by oxygen and Trolox with minimized bleaching via the oxygen radical scavenger cysteamine. Using this approach we resolved the single-molecule microsecond conformational fluctuations of two proteins: the two-state folder alpha-spectrin SH3 domain and the ultrafast downhill folder BBL.