Inverted multiphoton microscope

斯图加特大学细胞生物学和免疫学研究所分子肿瘤细胞生物学系的工作旨在研究癌症发生和癌症进展背景下的细胞转化过程。申请者对受体酪氨酸激酶生物学功能的基础研究为个性化癌症药物靶向方法的发展做出了重要贡献。然而，并不是所有的患者对这些靶向治疗都有反应，即使在最初的治疗中取得了成功，也往往会产生耐药性。为了开发有效的个体化肿瘤治疗策略，可靠的临床前模型系统是必不可少的，一方面在基础研究的背景下阐明分子关系，另一方面在应用研究中为药物筛选提供有效的平台。尽管建立了癌细胞系和小鼠肿瘤模型的经典2D细胞培养在复制肿瘤生理学方面都显示出相当大的局限性，但基于球体、器官和原代组织的异种3D培养可以用来以更现实的方式概括肿瘤的复杂性。因此，直接使用这种复杂的3D模型系统执行的分析保证了所获得的数据对患者的更高的可传递性。出于这个原因，申请人未来的工作将越来越多地使用体外(直接从生物体获得)和从头(合成重建)3D肿瘤组织培养，其准备工作将在斯图加特大学新成立的3R组织实验室进行。对于高细胞密度的组织样样品的实时检查，多光子显微镜特别适合，因为它的荧光成像深度可达一毫米。因此，需要一种具有集成的柔性多通道多光子成像的共焦显微镜，并具有额外的基于荧光寿命的能力。该显微镜将支持集成灌流系统以进行“液体处理”，并将装备用于在生理条件下对活组织样本进行长期观察。该仪器将集中放置在斯图加特研究中心系统生物学(SRCSB)的核心设施“细胞分析”中进行操作。一方面，这确保了最佳的技术支持；同时，将向SRCSB的所有工作组提供显微镜，从而向广大用户群体提供显微镜。购买该显微镜将大大扩展现场成像技术的范围，并将首次为斯图加特大学的研究人员提供多光子显微镜和基于荧光寿命的测量。

The work of the Department of Molecular Tumor Cell Biology at the Institute of Cell Biology and Immunology at the University of Stuttgart aims to investigate cellular transformation processes in the context of carcinogenesis and cancer progression. Fundamental studies by the applicant on the biological function of receptor tyrosine kinases have made an important contribution to the development of targeted approaches in personalized cancer medicine. However, not all patients respond to these targeted therapies and even with initial therapeutic success, resistance often develops. In order to develop effective personalized strategies for tumor therapy, reliable preclinical model systems are indispensable, on the one hand to elucidate molecular relationships in the context of basic research, and on the other hand to provide a valid platform for drug screening in applied studies. While both classical 2D cell cultures with established cancer cell lines and tumor models in mice show considerable limitations in replicating tumor physiology, heterogeneous 3D cultures based on spheroids, organoids and primary tissues can be used to recapitulate the complexity of tumors in a much more realistic manner. Analyses performed directly with such complex 3D model systems thus promise a higher transferability of the obtained data to the patient. For this reason, future work of the applicant will increasingly use ex vivo (directly obtained from the organism) and de novo (synthetically recreated) 3D tumor tissue cultures, the preparation of which will take place in the newly established 3R-Tissue Lab at the University of Stuttgart. For real-time examination of tissue-like samples of high cell density, multiphoton microscopy is particularly well suited due to its fluorescence imaging at penetration depths up to one millimeter. Therefore, a confocal microscope with integrated flexible multichannel multiphoton imaging is requested, with additional fluorescence lifetime-based capabilities. The microscope will support the integration of perfusion systems for "liquid handling" and will be equipped for long-term observations of living tissue samples under physiological conditions. The instrument will be centrally located and operated in the Core Facility "Cellular Analytics" of the Stuttgart Research Center Systems Biology (SRCSB). On the one hand, this ensures optimal technical support; at the same time, the microscope will be made available to all working groups of the SRCSB and thus to a broad user community. The acquisition of the microscope will significantly expand the range of imaging techniques on site and will for the first time make multiphoton microscopy and fluorescence lifetime-based measurements available to researchers at the University of Stuttgart.