Chameleon Sample Preparation Device for Cryo-EM

项目摘要 低温电子显微镜（cryo-EM）的最新技术进展预示着“分辨率 革命”，使高分辨率的结构，以确定各种目标， 避开了其他结构技术。尽管有这些进步，冷冻EM网格制备仍然是一个艰巨的任务。 在许多情况下，成功确定结构的障碍。事实上，尽管电子光学有所改进， 照相机技术和图像分析，以及对自动化的普遍推动，冷冻EM网格制备 仍然是一个费力且不可预测的过程，需要大量的人工用户输入来进行优化。的 加州大学圣地亚哥分校（UCSD）建立了世界一流的冷冻电镜设施， 电子显微镜和工作人员，但校园内的许多项目仍然停滞不前，由于无法 从纯化的标本制备高质量的冷冻EM网格。为了解决冷冻EM工作流程中的这一瓶颈， 在加州大学圣地亚哥分校，我们建议购买一个SPT实验室变色龙样品玻璃化仪器。本文书 代表了30多年来冷冻EM网格制备的第一次重大创新，采用了一种全新的方法 样品应用，产生更多可重复的冷冻EM网格，改善颗粒质量和分布， 通常增加了高分辨率结构确定的可能性。该仪器将集中在 在最近翻新的UCSD Cryo-EM设施内住宿和运营，并将提供给所有受过培训的 用户在加州大学圣地亚哥分校，周围的机构在拉霍亚“梅萨”地区，并在整个加州。对于UCSD的用户来说， 该工具将促进NIH资助的一系列生物医学重要项目的进展，这些项目包括 神经变性，病毒学，膜运输，DNA复制和修复，免疫学和蛋白质 生物成因和折叠

PROJECT SUMMARY Recent technological advances in cryogenic-electron microscopy (cryo-EM) have heralded a “resolution revolution” that has made it possible for high-resolution structures to be determined for a variety of targets that had eluded other structural techniques. Despite these advances, cryo-EM grid preparation remains a formidable roadblock to successful structure determination in many cases. Indeed, despite improvements in electron optics, camera technology, and image analysis, and a general drive towards automation, cryo-EM grid preparation remains a laborious and unpredictable process that requires extensive manual user input for optimization. The University of California, San Diego (UCSD) has established a world-class cryo-EM facility with state-of-the-art electron microscopes and staff, yet numerous projects across campus remain stalled due to an inability to prepare high-quality cryo-EM grids from purified specimens. To address this bottleneck in the cryo-EM workflow at UCSD, we propose to purchase an SPT Labtech Chameleon sample vitrification instrument. This instrument represents the first major innovation in cryo-EM grid preparation in over 30 years, with an entirely novel approach to sample application that yields more reproducible cryo-EM grids, improves particle quality and distribution, and generally increases the likelihood of high-resolution structure determination. The instrument will be centrally housed and operated within the recently renovated UCSD Cryo-EM Facility and will be available to all trained users at UCSD, surrounding institutions in the La Jolla “Mesa” area, and across California. For users at UCSD, this instrument will catalyze progress in a broad range of NIH-funded, biomedically-important projects spanning neurodegeneration, virology, membrane trafficking, DNA replication and repair, immunology, and protein biogenesis and folding.