A plasma focused ion beam microscope for Structural Cell Biology at the Astbury Biostructure Laboratory

结构生物学在生物学发现中是一个极其强大的工具。它可以揭示蛋白质和其他分子如何相互作用的分子细节，从而帮助我们了解在人/动物健康和福祉、食品安全和生物技术等领域具有巨大影响的基本生物过程。现代结构生物学的基石之一是一种名为冷冻电子显微镜的技术，这种技术使用强大的显微镜对蛋白质等生物材料拍摄高度放大的图像，从中我们可以计算出蛋白质复合体的三维结构。冷冻样品成像技术现在可以解决前所未有的规模和复杂性的结构问题，其结果正在改变我们对生物学的理解。自从最先进的泰坦克里奥斯显微镜安装在阿斯特伯里生物结构实验室(2016)，我们已经变得非常熟练地解决这些类型的结构，我们的社区，无论是利兹还是其他地方，已经解决了100‘S的高分辨率结构。这种方法取得了巨大的成功和影响，虽然这些结构本身具有巨大的信息量，但越来越明显的是，只有当我们能够理解这些结构在细胞和组织中的自然环境时，结构生物学的真正力量才能被释放。这一应用是通过购买一种名为等离子体聚焦离子束(PFIB)扫描电子显微镜(SEM)的新仪器来为ABSL带来新的能力。这台机器还将有一个集成在pFIB中的荧光显微镜。PFIB将通过以下方式帮助实现原位结构生物学工作流程的潜力：(A)使用我们可以引入的荧光标记物识别细胞和组织中感兴趣的区域，(B)在细胞和组织中铣削包含这些荧光标记物的薄片(非常薄的切片)，用于冷冻EM研究，以及(C)采用一种不同的技术，称为植物/动物组织的块面铣削。有了我们在ABSL现有的最先进的Titan Krios EM，这将使我们能够了解从毫米到纳米的长度尺度的生物学，改变Astbury生物结构实验室的结构发现，并为英国生物科学提供重要资源。

Structural biology is an enormously powerful tool in biological discovery. It can reveal the molecular details of how proteins and other molecules interact, and so help us understand fundamental biological processes that have enormous implications in areas as diverse as human/animal health and wellbeing, food security and biotechnology. One of the cornerstones of modern structural biology is a technique called cryo-electron microscopy, where powerful microscopes are used to take highly magnified images of biological material such as proteins, from which we can work out the three-dimensional structure of the protein complexes. CryoEM (imaging frozen samples) can now address structural problems of unprecedented scale and complexity, and the results are transforming our understanding of biology.Since the state of the art Titan Krios microscopes were installed in the Astbury Biostructure Laboratory (2016), we have become very adept at solving these type of structures, and our community, both in Leeds and beyond, has solved 100's of high-resolution structures. This approach has been enormously successful and impactful, and while these structures are immensely informative in their own right, it is becoming increasingly clear that the true power of structural biology will only be unlocked when we are able to understand these structures in their native contexts, within cells and tissues. This application is to bring a new capability to ABSL by purchasing a new instrument called a plasma focused ion beam (pFIB) scanning EM (SEM). This machine will additionally have a fluorescence microscope integrated into the pFIB. A pFIB will help realise the potential of in situ structural biology workflows by (a) identifying areas of interest in cells and tissues using fluorescent markers that we can introduce, (b) milling lamellae (very thin sections) that encompass these fluorescent markers, both in cells, and in tissue, for cryoEM studies, and (c) undertake a different technique called block face milling of plant/animal tissues. With our existing state-of-the-art Titan Krios EMs at ABSL, this will allow us to understand biology across lengths scales from millimetres to nanometers, transforming structural discovery at the Astbury Biostructure Laboratory, and providing a vital resource for UK bioscience.