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首页> 外文期刊>Journal of the Royal Society Interface >Nanometric depth resolution from multi-focal images in microscopy
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Nanometric depth resolution from multi-focal images in microscopy

机译：显微镜中多焦点图像的纳米深度分辨率

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摘要

We describe a method for tracking the position of small features in three dimensions from images recorded on a standard microscope with an inexpensive attachment between the microscope and the camera. The depth-measurement accuracy of this method is tested experimentally on a wide-field, inverted microscope and is shown to give approximately 8 nm depth resolution, over a specimen depth of approximately 6 μm, when using a 12-bit charge-coupled device (CCD) camera and very bright but unresolved particles. To assess low-flux limitations a theoretical model is used to derive an analytical expression for the minimum variance bound. The approximations used in the analytical treatment are tested using numerical simulations. It is concluded that approximately 14 nm depth resolution is achievable with flux levels available when tracking fluorescent sources in three dimensions in live-cell biology and that the method is suitable for three-dimensional photo-activated localization microscopy resolution. Sub-nanometre resolution could be achieved with photon-counting techniques at high flux levels.
机译：我们描述了一种方法，用于从标准显微镜上记录的图像以三维方式跟踪小特征的位置，而显微镜和相机之间的连接较为便宜。该方法的深度测量精度已在宽视野倒置显微镜上进行了实验测试，结果表明，当使用12位电荷耦合器件时，在大约6μm的样品深度下，该方法的深度分辨率约为8 nm（ CCD）相机和非常明亮但尚未分解的颗粒。为了评估低通量限制，使用理论模型来导出最小方差界限的解析表达式。使用数值模拟测试分析处理中使用的近似值。结论是，当在活细胞生物学中以三维方式跟踪荧光源时，利用可用的通量水平可获得约14 nm的深度分辨率，并且该方法适用于三维光激活定位显微镜分辨率。可以通过在高通量水平下使用光子计数技术来实现亚纳米分辨率。

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