Computed Tomography using projection X-ray microscope for Biological materials

投影x射线显微镜利用扫描电子显微镜（SEM）的聚焦电子束激发的薄（0.1-3 μm）目标金属膜上的非常小的x射线源。如果一个物体被放置在目标金属膜的正下方，发散的x射线会放大物体的阴影。由于x射线光源的尺寸小于100nm，因此该显微镜的分辨率约为0.1μm，优于光学显微镜。我们已将该显微镜应用于三维结构分析，通过锥形束计算机断层扫描。在步进电机控制的样品旋转器上，记录小昆虫Omiscus Porcellio全方向（360°）每1°的投影图像。采用临界点干燥法制备Omiscus Porcellio，并将其固定在样品旋转器的0.4μm厚的薄胶带上。图像用直接探测x射线的冷却CCD相机（HAMAMATSU C4880）记录。利用反投影算法对玉米梁投影进行三维图像重建。重建的三维图像（256 × 256 × 256）分辨率约为3 μm。消化器官的三维图像清晰可见。我们将分析更小的对象，比如包含更详细结构的神经网络。

The projection X-ray microscope makes use of a very small X-ray source from a thin (0.1-3 μm) target metal film that is excited by the focused electron beam of a scanning electron microscope (SEM). If an object is placed just below the target metal film, the diverging X-rays enlarge the shadow of the object. Because the size of X-ray light source is less than 100nm, the resolution of this microscope is about 0.1μm which is better than that of optical microscopes. We have applied this microscope for three-dimensional (3-D) structure analysis by means of cone-beam computerized tomography. The projection images of small insect, Omiscus Porcellio, were recorded in every 1° for whole direction (360°) with a stepping motor controlled sample rotator. An Omiscus Porcellio was prepared by critical point drying method and fixed on a thin adhesive tape (0.4μm thick) of the sample rotator. The images were recorded with cooled CCD camera (HAMAMATSU C4880) which detect X-ray directly. The 3-D image was reconstructed from corn-beam projections using back-projection algorithm. The resolution of the reconstructed 3-D image (256 x 256 x 256) was about 3 μm. The digestive organs were clearly visualized in 3-D.We are going to analyze smaller objects such as nerve networks which contains more detailed structure.