Automated Microscope for Long-Duration, Quantitative Dynamic Imaging

描述（由申请人提供）：我们请求资金购买Perkin Elmer Opera LX显微镜，用于全自动点扫描共聚焦高通量微孔板成像。范德比尔特大学的传统倒置和共聚焦显微镜、读板仪和高通量扫描仪无法支持对动态细胞事件的自动化长期、高分辨率观察，例如大量单个细胞中的趋化作用、趋化性和趋触性、细胞分裂和增殖、细胞器功能、受体和蛋白质运输以及凋亡。在范德比尔特没有乐器可以与Opera LX相媲美。Opera LX提供的主要功能是微透镜旋转圆盘共聚焦显微镜，配有10倍空气物镜和20倍和60倍自动水物镜，三个激光器和一个可调紫外光源，红外自动对焦，两个摄像头，可同时进行双色成像和顺序双色成像，以及快速插入和XYZ定位板。双核处理器允许多个用户同时进行图像采集和复杂处理。最重要的是，Opera可以在数小时至数天内并行观察维持在96或384孔板或定制微流体装置中的数百个单独的活细胞群体的细胞行为变化。该仪器将支持来自文理学院和医学与工程学院13个部门的12个主要用户和14个次要用户的高度协作组。主要用户正在进行的NIH资助的项目将受益，包括对5-羟色胺蛋白转运蛋白运输的研究;结构化微环境中的细胞行为，包括趋化性、触变性、旁分泌信号和内皮附着;基因组维持蛋白的鉴定和表征，调节PIP 3产生的基因，以及ENOX 1在暴露于辐射的内皮细胞形态发生中的作用;以及干细胞介导心脏缺血性损伤的能力。次要项目包括受体运输，信号动力学，心脏肌节周转，细胞基质相互作用，以及细菌和病毒的感染性。对于每个项目，所要求的仪器将通过提供所研究的生物现象的独特的新测量能力而产生直接和实质性的影响。范德比尔特综合生物系统研究与教育研究所（VIIBRE）对该显微镜的采购和操作将保证用户培训/支持，24小时/天7天/周的访问以及仪器维护和维修。用户费用将包括工作人员的支持，服务和维修。其他研究人员将能够利用可用的时间。VIIBRE在开发和提供先进的微流体和微加工技术、电子设备、培训和用户设施方面有着出色的业绩，范德比尔特的200多名学生和教师都有着这样的业绩。公共卫生关系万斯：拟议的最先进的仪器将为研究人员提供新的能力，以研究涉及复杂动态细胞内和细胞外信号网络的发育，伤口修复，衰老，癌症和心力衰竭过程，从而提高我们保持健康和诊断和治疗疾病的能力。

DESCRIPTION (provided by applicant): We request funds to purchase a Perkin Elmer Opera LX microscope for fully automated, point-scanning confocal, high-throughput microplate imaging. Conventional inverted and confocal microscopes, plate readers and high-throughput scanners at Vanderbilt University cannot support automated long-term, high-resolution obser- vation of dynamic cellular events such as chemokinesis, chemotaxis and haptotaxis, cell division and differen- tiation, organelle function, receptor and protein trafficking, and apoptosis in large numbers of individual cells. There are no instruments at Vanderbilt comparable to the Opera LX. Key features offered by the Opera LX are the microlens spinning disk confocal microscope with a 10x air objective and 20x and 60x automated water objectives, three lasers and a tunable UV source, IR autofocus, two cameras that enable simultaneous two-color imaging and sequential multicolor imaging, and rapid insertion and XYZ positioning of plates. The dual-core processor allows simultaneous image acquisition and sophisticated processing by multiple users. Most importantly, the Opera can observe in parallel the changes in cellular behavior over hours to days for hundreds of separate populations of live cells maintained in 96 or 384 well plates or custom-fabricated microfluidic devices. This instrument will support a highly collaborative group of 12 major and 14 minor users from 13 departments in the College of Arts & Science and the Schools of Medicine and Engineering. Ongoing, NIH-funded projects by major users that will benefit include research on serotonin protein transporter trafficking; cellular behavior including chemotaxis, haptotaxis, paracrine signaling, and endothelial attachment in structured microenviron- ments; identification and characterization of genome maintenance proteins, the genes that regulate PIP3 pro- duction, and the role of ENOX1 in the morphogenesis of endothelial cells exposed to radiation; and the ability of stem cells to mediate cardiac ischemic injury. Minor projects include receptor trafficking, signaling dynamics, cardiac sarcomere turnover, cell-matrix interactions, and bacterial and viral infectivity. For each project, the requested instrumentation will have an immediate and substantial impact by providing unique new measurement capabilities of the biological phenomena under study. Acquisition and operation of this microscope by the Vanderbilt Institute for Integrative Biosystems Research and Education (VIIBRE) will guarantee user training/support, 24 hour/day 7 day/week access, and instrument maintenance and repair. User fees will cover staff support, service and repair. Other reseachers will be able to use available time. VIIBRE has an outstanding re- cord in developing and delivering advanced microfluidic and microfabrication technologies, electronic instru- mentation, training, and user facilities to over 200 students and faculty at Vanderbilt. Public Health Rele- vance: The proposed state-of-the-art instrument will give researchers a new capability to study processes in development, wound repair, aging, cancer and heart failure that involve complex dynamic intracellular and extracellular signaling networks, thereby improving our ability to maintain health and diagnose and treat disease.