Energy-Filtered Electron Tomography

我们进一步开发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300 kV场发射透射电子显微镜上实现，该显微镜配备了先进的成像滤波器，包括2048 x 2048像素CCD相机，具有高探测量子效率和快速读出。采集是通过灵活的计算机脚本来控制的，它可以校正标本漂移和连续倾斜角之间的散焦，并在每个倾斜角的定义能量下收集能量过滤图像。投影元素分布是通过获取能量损失谱中特征核心边缘上方和下方的图像，并减去每个像素的外推背景强度来获得的。我们实现了一种双轴同步迭代重建技术（SIRT），与传统加权傅立叶反投影重建相比，该技术可以提高信噪比。通过应用考虑复数非弹性散射的厚度校正算法，并结合核壳电子激发的散射截面，我们已经表明，可以根据每体素的原子数来量化元素分布。目前正在进行的实验是对磷进行三维成像，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.