Local dimensionality determines imaging speed in localization microscopy.
Local dimensionality determines imaging speed in localization microscopy.
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DOI：
10.1038/ncomms13558
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发表时间：
2017-01-12
影响因子：
16.6
通讯作者：
Cox S
中科院分区：
文献类型：
--
作者：
Fox-Roberts P;Marsh R;Pfisterer K;Jayo A;Parsons M;Cox S
定位显微镜允许生物样品以数十纳米的长度尺度成像。活细胞超分辨率成像是罕见的，因为它通常被认为是太慢的动态样本。数据采集的速度可以通过调整每个时间帧中激活的荧光团的密度来优化。在这里，我们表明，特定结构的最大可实现的成像速度按数量级变化，这取决于样品的维度（即，样品是否更像一个点，一股或一个扩展的结构，如焦点粘合）。如果使用太高的激励密度，我们证明，分析经历无声的失败，导致重建文物。我们正在发布一个工具，允许用户在活细胞和固定细胞实验中识别图像中激活密度过高的区域并对其进行校正。局部化显微镜能够对生物样品进行纳米级成像，但该方法太慢，无法用于动态系统。在这里，作者开发了一个数学模型，优化了所需的帧数，并估计了超分辨率成像的最大速度。
Localization microscopy allows biological samples to be imaged at a length scale of tens of nanometres. Live-cell super-resolution imaging is rare, as it is generally assumed to be too slow for dynamic samples. The speed of data acquisition can be optimized by tuning the density of activated fluorophores in each time frame. Here, we show that the maximum achievable imaging speed for a particular structure varies by orders of magnitude, depending on the sample dimensionality (that is, whether the sample is more like a point, a strand or an extended structure such as a focal adhesion). If too high an excitation density is used, we demonstrate that the analysis undergoes silent failure, resulting in reconstruction artefacts. We are releasing a tool to allow users to identify areas of the image in which the activation density was too high and correct for them, in both live- and fixed-cell experiments. Localisation microscopy enables nanometre-scale imaging of biological samples, but the method is too slow to use on dynamic systems. Here, the authors develop a mathematical model that optimises the number of frames required and estimates the maximum speed for super-resolution imaging.