Energy-Filtered Electron Tomography

我们进一步发展了一种基于能量过滤电子断层扫描的新技术，用于成像细胞中特定化学元素的三维分布。该技术已在300KV场发射电子显微镜上实现，该显微镜配有先进的成像滤光片，包括一台探测量子效率高、读出速度快的2048×2048像素CCD相机。采集通过灵活的计算机脚本进行控制，可以对连续倾斜角度之间的样品漂移和散焦进行校正，并以每个倾斜角度的定义能量收集经过能量过滤的图像。通过获取能量损失谱中特征核心边缘上下的图像并减去每个像素的外推背景强度来获得投影的元素分布。我们实现了一种双轴同时迭代重建技术(SIRT)，与传统的加权傅立叶反投影重建相比，它可以提高信噪比。通过应用考虑多次非弹性散射的厚度校正算法，以及结合核壳电子激发的散射截面，我们已经证明了用每个体素的原子数量来量化元素分布是可能的。对磷进行三维成像的实验正在进行中，从而确定DNA在细胞核染色质中的分布。

We have developed further a new technique based on energy-filtered electron tomography for imaging the three-dimensional distribution of specific chemical elements in cells. The technique has been implemented in a 300 kV field-emission transmission electron microscope equipped with an advanced imaging filter including a 2048 x 2048 pixel CCD camera with high detective quantum efficiency and fast read-out. Acquisition is controlled by means of flexible computer scripts, which enable correction for specimen drift and defocus between successive tilt angles and collection of energy-filtered images at defined energy for each tilt angle. Projected elemental distributions are obtained by acquiring images above and below characteristic core-edges in the energy-loss spectrum and by subtracting the extrapolated background intensity at each pixel. We have implemented a dual-axis simultaneous iterative reconstruction technique (SIRT) to improve the signal-to-noise ratio compared with that obtainable by a conventional weighted Fourier back-projection reconstruction. By applying a thickness correction algorithm that takes into account plural inelastic scattering, and by incorporating scattering cross sections for excitation of core-shell electrons, we have shown that it is possible to quantify the elemental distributions in terms of the number of atoms per voxel. Experiments are in progress to image phosphorus in three dimensions and hence to determine the distribution of DNA in the chromatin of the cell nucleus.