Soft X-ray Nanotomography of Cryo Specimens

为了了解生物标本中结构之间的空间关系，需要三维成像。软x射线断层扫描提供了以30-100纳米体素分辨率成像几微米厚标本的可能性，独特的对比机制，以及检查水合标本的能力。采用软x射线光学技术的纳米层析成像技术将在带有液氮温度台的软x射线扫描透射显微镜上增加层析成像能力。菲涅尔带片将软x射线波动光束聚焦到一个30-50纳米分辨率的探针（任何波长的电磁辐射的最小聚焦点），并通过该探针在一定倾斜角度范围内扫描样品。景深是这样的，每个图像将代表一个投影通过标本，至少在为初步研究选择的问题。在对冻干标本进行初步研究后，玻璃化的水化标本将在液氮温度下成像。这些新颖的纳米层析成像能力将与我们的生物学合作者在我们的x射线显微镜用户设施中用于正在进行的新的跨学科研究。这些合作包括中期染色体染色质折叠的研究，疟原虫恶性疟原虫的成像，蛋白质和DNA浓度的定量制图，以了解精子中的染色体堆积，软骨和骨的钙化（对关节炎和骨质疏松症的影响），以及细胞核中RNA转录位点的定位。

To understand the spatial relationships among structures within biological specimens, three-dimensional imaging is required. Soft x-ray tomography offers the possibility to image several micrometer thick specimens at 30-100 nanometer voxel resolution, unique contrast mechanisms, and the ability to examine hydrated specimens. Nanotomography with soft x-ray optics will be developed by adding tomography capabilities to a soft x-ray scanning transmission microscope with a liquid nitrogen temperature stage. Fresnel zone plates will be used to focus a soft x-ray undulator beam to a 30-50 nanometer resolution probe (the smallest focused spot of electromagnetic radiation of any wavelength), and the specimen will be scanned through this probe at a range of tilt angles. The depth of field is such that each image will represent a projection through the specimen, at least in problems selected for initial studies. After initial studies of freeze-dried specimens, hydrated specimens which have been vitrified will be imaged at liquid nitrogen temperatures. These novel nanotomography capabilities will be employed in ongoing and new interdisciplinary studies with our biology collaborators at our x-ray microscope user facility. These collaborations have included the study of chromatin folding in metaphase chromosomes, imaging of the malarial parasite P. falciparum, quantitative mapping of protein and DNA concentrations to understand chromosome packing in sperm, calcification in cartilage and bone (with implications for arthritis and osteoporosis), and the localization of RNA transcription sites in the nucleus.