Lightsheet-Microscopy-System

为马格德堡大学医学院的跨学科研究平台“多参数生物成像和细胞学”(MPBIC)提出了一种白光激光光片显微镜。MPBIC平台是医学院系中光学显微镜和流式细胞术方法的中心研究基础设施，其研究重点是“炎症的免疫学和分子医学”。MPBIC目前可用的成像技术(多光子显微镜、共焦显微镜、广域显微镜)允许以高分辨率显示小样本或大样本的小切片。由于激发光或发射光的穿透和发射深度受限，较大的样品必须切割成薄片，这会导致制备伪影和样品损失。然而，不同的研究目标要求在完整的三维组织背景下分析细胞渗透或组织适应不良/结构变化，这不能通过薄片(或仅是不完全的)来解决。多光子活体显微镜是MPBIC平台中的一种成熟程序，它允许分析三维组织中的细胞动力学，但仅限于小样本区域，尽管激发激光的穿透深度很高。因此，这种方法仅对解决炎症过程的研究项目有限有用，炎症过程不是均匀分散的，而是以渗透或感染点的形式发生的。最近，已经发展了各种整体染色和随后的组织清除的方法，使得能够用荧光标记的抗体检测完整器官中的结构。在实践中，由于清除与MPBIC中可用的扫描技术相结合导致了极长的测量时间，因此该程序不适合在教师中广泛应用。因此，用目前的显微镜或数量非常有限的显微镜，不可能对完整的器官或组织进行可视化和分析。这里提出的光片显微镜将满足教职员工对从宏观到微观的大而完整的生物样本(实验动物的器官、器官和人体活检组织)进行快速3D分析的日益增长的需求。这将能够检测到组织水平上的解剖结构变化/修改，直到免疫细胞的渗透，或者不均匀分散的炎症中的结构变化，或者具有细胞-亚细胞分辨率的感染点，这对于许多有前途的正在进行和计划中的研究项目来说是必不可少的。

A white light laser Lightsheet microscope is proposed for the interdisciplinary research platform "Multiparametric Bioimaging and Cytometry" (MPBIC) of the Medical Faculty of the Otto-von-Guericke-Universität Magdeburg. The MPBIC platform is the central research infrastructure for light microscopy and flow cytometry approaches in the medical faculty within the research focus "Immunology and molecular medicine of inflammation". The imaging techniques currently available in the MPBIC (multiphoton-microscopy, confocal microscopy, wide-field microscopy) allow for the visualization of small samples or small sections of larger samples with high resolution. Larger samples have to be cut into thin sections due to the restricted penetration and emission depth of the exciting or the emitted light, which is leading to preparation artefacts and sample loss. However, various research objectives require the analyses of cell infiltrates or tissue maladaptation/structural alterations in the intact three-dimensional tissue context, which can be not addressed with thin sections (or only incompletely). The multi-photon intravital microscopy, which is a well-established procedure in the MPBIC platform, allows for the analysis of cell dynamics in the three-dimensional tissue but is limited to small sample regions despite the high penetration depth of the exciting laser. Consequently, this method is only limitedly useful for research projects addressing inflammatory processes, which are not homogeneously dispersed, but occurring in form of infiltration or infection spots. Recently, various methods of whole-mount staining and subsequent tissue clearing have been developed that enable the detection of structures in whole intact organs with fluorochrom-conjugated antibodies. Since, in practice, the combination of clearing with the scanning techniques available in the MPBIC leads to extremely long measuring times, this procedure is unsuitable for a broad application in the faculty. Therefore, the visualization and analysis of whole intact organs or tissues is not possible with the currently present microscopes or in a very restricted number. The herein proposed lightsheet-microscope will cover the increasing need in the faculty for fast 3D-analyses of large and intact biological samples (organs of experimental animals, organoids, and human biopsies) from the macro into the micro level. This will enable the detection of anatomical-structurally alterations/modifications in the tissue level up to infiltrations of immune cells, or structural alterations in inhomogeneously dispersed inflammation- or infection spots with cellular-subcellular resolution, which is indispensable for a number of promising running and planned research projects.