Advancing Multi-Color EM via Direct Detector-enabled 4D-STEM

通过支持直接检测器的 4D-STEM 推进多色 EM

• 批准号: 10031737
• 负责人: Mark H Ellisman
• 金额: $ 36.51万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10031737
• 关键词: 3-Dimensional; Advanced Development; Algorithms; Antibodies; Big Data; Biological; Cell Nucleus; Cells; Characteristics; Chemicals; Color; Complex; Computer Analysis; Computer software; Computing Methodologies; Cryoelectron Microscopy; Data Set; Detection; Development; Devices; Dimensions; Dose; Dose-Rate; Electron Beam; Electron Microscopy; Electron energy loss spectroscopy; Electrons; Elements; Energy-Filtering Transmission Electron Microscopy; Feasibility Studies; Gold Colloid; Heart; Image; Imaging technology; Label; Laboratories; Measures; Metals; Methodology; Methods; Modernization; Molecular; Molecular Structure; Monte Carlo Method; Nature; Noise; Osmium; Outcome; Performance; Positioning Attribute; Preparation; Process; Quantum Dots; Resolution; Roentgen Rays; Sampling; Scanning; Signal Transduction; Specimen; Speed; Stains; Technology; Testing; Thick; Tissues; Transmission Electron Microscopy; Validation; Visualization; Work; base; computerized data processing; contrast imaging; detector; flexibility; improved; industry partner; interest; light microscopy; materials science; metal chelator; microscopic imaging; new technology; next generation; novel; sensor technology; simulation; software development; structural biology; technology development

PROJECT SUMMARY This focused technology development project will advance into first practice a transformative new imaging technology for multicolor electron microscopy (ColorEM), enabling new capabilities for high resolution and 3 dimensional (3D) localization and differentiation of different molecular complexes in cells and tissues. With this technology, we will substantively improve upon presently practiced, multiple labeling strategies involving antibody labelling approaches detected using colloidal gold, quantum dots, etc.. We will further provide a more accessible, higher throughput, and dose efficient alternative to spectroscopic methods, like energy dispersive x-ray (EDX) and electron energy loss spectroscopy (EELS) / energy filtered transmission electron microscopy (EFTEM). At the heart of this effort, we will exploit the performance characteristics of a next- generation, ultra-high speed direct detection device (DDD), now being optimized and extended (by us with an industry partner) for use with scanning transmission electron microscopy (STEM), a broadly deployed capability on modern EM columns. The optimized application of this new pixelated STEM detector technology, in combination with an extended palette of elemental probes and advanced software analysis, will allow for unprecedented new capabilities for performing atomic number (Z) contrast imaging – a process we call ColorSTEM. To bring this new integrated methodology into first practice, we will build on recent feasibility studies and surmount remaining technical hurdles to 1) make multi-labelled EM specimens with optimized elemental probe combinations; 2) troubleshoot the first use of a potentially transformative new sensor technology and work out the methodology for employing it for Z contrast imaging; and 3) devise the associated analytical/computational methodology for processing the very large resultant data sets to enable unambiguous differentiation of molecular complexes selectively marked with elemental probes.

项目摘要 这个重点的技术开发项目将晋升为首次练习一种变革性的新成像 多色电子显微镜技术（COLOREM），为高分辨率和3 细胞和组织中不同分子复合物的尺寸（3D）定位和分化。与此 技术，我们将对目前正在实践的多个标签策略进行实质性改进 使用胶体金，量子点等检测到的抗体标记方法。我们将进一步提供更多 可访问，更高的吞吐量和剂量有效替代光谱法，例如能量色散 X射线（EDX）和电子能量损失光谱（EEL） /能量过滤透射电子 显微镜（EFTEM）。这是这项努力的核心，我们将利用下一步的性能特征 一代，超高速度直接检测装置（DDD），现在已进行了优化和扩展（我们是由我们 行业合作伙伴）用于扫描传输电子显微镜（STEM），这是一个广泛部署的 现代EM柱上的能力。该新像素化的茎检测器的优化应用 技术，结合了元素问题和高级软件分析的扩展调色板， 将允许执行原子数（z）对比成像的前所未有的新功能 - a 流程我们称之为色彩系统。为了将这种新的综合方法成为第一练习，我们将基础 最近的可行性研究和超越剩余的技术障碍到1）制作多标签的EM标本 具有优化的元素探针组合； 2）对潜在的变革新的新用途进行故障排除 传感器技术并确定将其用于Z对比成像的方法； 3）设计 相关的分析/计算方法，用于处理非常大的结果数据集以启用 分子复合物的明确分化选择性地标记了元素问题。