High Resolution 3D X-ray Diffraction Microscope

高分辨率 3D X 射线衍射显微镜

• 批准号: 7178506
• 负责人: Chris Johnson Jacobsen
• 金额: $ 27.42万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-08-01 至 2009-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7178506
• 关键词: Resolution; 3D; ray; Diffraction; Microscope

DESCRIPTION (provided by applicant): We propose to develop x-ray diffraction microscopy towards the goal of 10 nm resolution 3D imaging of frozen hydrated cells. This goal is built upon our recent success in obtaining 30 nm resolution 2D images of a freeze-dried yeast, Saccharomyces cerevisiae, in experiments carried out at the Advanced Light Source (ALS) at Lawrence Berkeley National Laboratory. X-ray diffraction microscopy is a-recently-developed method in which the coherent diffraction pattern of a non-crystalline sample is measured, and then phased by an iterative algorithm to yield a real-space image. It provides an approach to achieve the maximum possible spatial resolution in x-ray imaging by avoiding the radiation dose increasing limitations of optic efficiency and maximum collection angle otherwise imposed by an x-ray optical system. With NIH support, we have built a new apparatus to acquire tilt series data of frozen hydrated specimens, and have installed it on a beam line at the ALS. With this apparatus, we have collected a series of 2D diffraction patterns of freeze-dried yeast, and have obtained reconstructed images at 30 nm resolution. We have also begun to collect diffraction data from frozen hydrated yeast, and have verified that the radiation tolerance of frozen hydrated cells is compatible with our goal of 10 nm resolution. To reach our goal of high resolution 3D imaging of frozen hydrated specimens, we plan on providing specimen preparation capabilities near the apparatus, and to improve our apparatus to allow for more rapid evaluation of different preparations by incorporating a scanned image capability. We also plan on improving our data acquisition procedures by developing a new beamstop to allow us to better capture data at low spatial frequencies. We will improve our data reconstruction approaches and software to deal with the challenges posed by 3D imaging. We will carry out labeling studies and correlative microscopy so as to interpret images of yeast delivered by this new modality. We will improve the beam line we presently operate at to provide the capability to work at higher photon energies so as to work with thicker specimens. We will also take steps toward the establishment of a new facility (including a custom designed undulator and beam line) at the Advanced Light Source that will make the technique routinely available to the wider user community. This project has top priority in the ALS Strategic Plan. Development of improved methods for studying cellular ultrastructure is crucial for increasing our understanding of biological structure and function. Soft x rays have fundamental advantages for imaging unsectioned, hydrated cells, and diffraction microscopy can maximize the structural information that can be obtained for a given radiation exposure.

描述（由申请人提供）：我们建议开发x射线衍射显微镜，以实现冷冻水合细胞10纳米分辨率3D成像的目标。这一目标是建立在我们最近在劳伦斯伯克利国家实验室的先进光源（ALS）进行的实验中成功获得30纳米分辨率的冷冻干酵母，Saccharomyces cerevisiae的2D图像。x射线衍射显微镜是最近发展起来的一种方法，其中测量非晶体样品的相干衍射模式，然后通过迭代算法进行相位处理以产生实空间图像。它提供了一种在x射线成像中实现最大可能空间分辨率的方法，避免了辐射剂量增加对光学效率和最大收集角的限制，否则由x射线光学系统施加。在美国国立卫生研究院的支持下，我们建立了一个新的设备来获取冷冻水合标本的倾斜序列数据，并将其安装在ALS的束流线上。利用该仪器，我们收集了冻干酵母的一系列二维衍射图，并获得了30 nm分辨率的重建图像。我们也开始收集冷冻水合酵母的衍射数据，并验证了冷冻水合细胞的辐射耐受性符合我们10纳米分辨率的目标。为了实现冷冻水合标本的高分辨率3D成像目标，我们计划在设备附近提供标本制备能力，并通过结合扫描图像能力来改进我们的设备，以便更快地评估不同的制剂。我们还计划通过开发一种新的波束阻挡器来改进我们的数据采集程序，使我们能够更好地捕获低空间频率的数据。我们将改进数据重建方法和软件，以应对3D成像带来的挑战。我们将进行标记研究和相关的显微镜，以便解释这种新方式传递的酵母图像。我们将改进我们目前操作的光束线，以提供在更高光子能量下工作的能力，以便处理更厚的样品。我们还将采取措施，在先进光源建立一个新的设施（包括定制设计的波动器和光束线），这将使该技术常规地提供给更广泛的用户社区。这个项目是ALS战略计划的重中之重。研究细胞超微结构的改进方法对于提高我们对生物结构和功能的认识至关重要。软x射线在成像未切片的水合细胞方面具有基本优势，而衍射显微镜可以最大限度地获得给定辐射暴露的结构信息。