Bayesian localization microscopy reveals nanoscale podosome dynamics.

Bayesian localization microscopy reveals nanoscale podosome dynamics.
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DOI:
10.1038/nmeth.1812
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发表时间:
2011-12-04
期刊:
影响因子:
48
通讯作者:
Heintzmann, Rainer
Heintzmann, Rainer
中科院分区:
生物学1区
文献类型:
--
作者:
Cox, Susan;Rosten, Edward;Monypenny, James;Jovanovic-Talisman, Tijana;Burnette, Dylan T.;Lippincott-Schwartz, Jennifer;Jones, Gareth E.;Heintzmann, Rainer

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我们展示了一种定位显微镜分析方法,该方法能够使用标准荧光蛋白和氙弧灯照明在活细胞中提取结果。我们的闪烁和漂白的贝叶斯分析(3B分析)方法将整个数据集同时建模为由许多荧光团生成,这些荧光团可能在任何给定时间发光,也可能不发光。由此产生的技术允许许多重叠的荧光团在每个帧中,并统一的分析从闪烁和漂白事件的定位。通过对整个数据集进行建模,我们能够使用荧光团的每次再现来提高定位精度。该技术的高性能使我们能够在4秒的时间尺度上以50 nm的分辨率揭示podosome形成和解离的纳米级动力学。
We demonstrate a localization microscopy analysis method that is able to extract results in live cells using standard fluorescent proteins and Xenon arc lamp illumination. Our Bayesian analysis of blinking and bleaching (3B analysis) method models the entire dataset simultaneously as being generated by a number of fluorophores which may or may not be emitting light at any given time. The resulting technique allows many overlapping fluorophores in each frame, and unifies the analysis of localization from blinking and bleaching events. By modeling the entire dataset we are able to use each reappearance of a fluorophore to improve the localization accuracy. The high performance of this technique allows us to reveal the nanoscale dynamics of podosome formation and dissociation with a resolution of 50 nm on a four second timescale.
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