Confocal spinning disk high throughput screening microscope

我们正在申请资金，以获得几个正在进行和计划中的跨学科研究项目中迫切需要的共焦成像细胞仪。高端免疫荧光显微镜的可能性和技术要求在过去几年中稳步扩大，我们目前在这一领域的需求无法再通过CharitéCampus Mitte的现有显微镜基础设施来满足(柏林Charité三个地点中的一个)。最近，这导致我们在许多项目中的显微镜实验显著延迟，也影响了我们能够生成的数据的质量。此外，由于现有显微镜的技术限制，我们无法建立一些要求更高的实验装置，这些装置将极大地受益于现有的项目，并需要建立新的研究方向。最近，显微镜的设计取得了巨大进展，生物学中出现了许多新的先进应用：更灵敏的相机、更大的视场和越来越完全自动化的图像采集工作流程，以及新的光学设计，使得在很长一段时间内从数百个样品中获取高质量的3D共焦图像成为可能。因此，系统地跟踪单个细胞在自然环境中的3D时间成为可能，这对于更好地了解许多与医学直接相关的生物过程是必不可少的：例如癌症的发展，组织内免疫反应的跟踪，但也有许多细胞生物学中的问题，例如需要在有机物中进行实验。成像细胞仪的另一个重要应用将是获得患者组织标本的高分辨率共聚焦图像，用新的多路复用方案进行荧光染色，使同一张载玻片能够连续染色数十种蛋白质。用校园现有的设备获取这种图像非常耗时，因此实际上是不可行的。随附的图像分析包还将允许我们以自动化的方式以高吞吐量对采集过程中的数据进行动态分析。总之，共聚焦成像细胞仪将确保提供用于高通量3D活细胞成像的尖端显微镜设备，以及适合量化如此大量数据的专用软件，并帮助我们建立基于显微镜的新分析方法。这也将使相关小组的成员更容易为现有项目获得足够的显微镜时间，而依靠校园现有设备进行现有项目已变得越来越困难。在随附的申请表中，我们包括了一份详细的计划，该计划将如何将细胞仪用作共同申请者的七个不同实验室的许多现有和计划中的研究项目。

We are applying for funding to acquire a confocal imaging cytometer that is urgently needed in several ongoing and planned interdisciplinary research projects.The possibilities, but also the technical requirements in high-end immunofluorescence microscopy have steadily expanded during the last few years and our current needs in this area cannot be met anymore with the available microscopy infrastructure on Charité Campus Mitte (one out of 3 Charité sites in Berlin). Recently, this has led to significant delays in our microscopy experiments in many projects and also affected the quality of the data we were able to generate. Furthermore, because of technical limitations of the available microscopes, we were not able to establish some more demanding experimental setups that would have greatly benefitted existing projects and are required to establish new research directions.Recently, enormous progress has been made in the design of microscopes and lots of new advanced applications in biology have emerged: more sensitive cameras, larger field of view and increasingly fully automated image acquisition workflows, as well as new optical designs make it possible to acquire high quality confocal images from hundreds of samples in 3D over long periods of time. Thus, it becomes possible to systematically track single cells in 3D over time in their natural environment, which is essential for a better understanding of many biological processes that are of immediate relevance in medicine: examples include the development of cancer, tracking of the immune response within tissues, but also many questions in cell biology that require for example experiments in organoids.Another important application for the imaging cytometer will be to obtain high-resolution confocal images of histological specimens from patients, fluorescently stained with new multiplexed protocols that enable sequential staining of the same slide for dozens of proteins. Acquisition of such images with the equipment currently available on campus is extremely time-consuming and therefore practically not feasible. The included image analysis package will also allow us to analyze the data on the fly during acquisition in an automated manner in high throughput.In summary, the confocal imaging cytometer will ensure the availability of cutting edge microscopy equipment for high-throughput 3D live-cell imaging coupled with dedicated software appropriate for quantification of such large amounts of data and help help us to establish new microscopy-based assays. It will also make it easier for members of the involved groups to get enough microscopy time for existing projects, which has become increasingly difficult relying on the existing equipment on campus. In the included application forms, we include a detailed plan how the cytometer will be used as benefit many existing and planned research projects of seven different labs that act as coapplicants.