Custom-built Digital Scanned Laser Light Sheet Microscope (DSLM)

描述(由申请人提供)：我们要求定制一台数字扫描激光光片显微镜(DSLM)，它将使芝加哥的一大批研究人员能够以高分辨率和高速度显示活组织和生物中的蛋白质，并且不会与其他荧光显微镜技术相关的样品损坏。一个主要的用户群体将是芝加哥系统生物学中心(CCSB)的研究人员(www.chicago-center-for-system-biology.org/；Grant P50GM081892)。该中心的科学计划侧重于生理、发育和进化尺度上转录网络的动态。新系统将特别使我们能够生成和分析发育、正常和疾病进展方面的表达网络的广泛文库。由于该仪器的独特功能，我们还收到了来自芝加哥大学以外的团体的请求，我们将为这些用户(小用户组)保留25%的使用该仪器的时间。DSLM是由欧洲分子生物学实验室(EMBL)的Stelzer博士团队开发的一种创新的实时成像荧光显微镜系统(Keller等人，2008)。科学，322：1065-9)。与其他先进的荧光显微镜技术(共焦和双光子)相比，DSLM提供了50多倍的成像速度和10倍的信噪比，同时将样品暴露在至少两个数量级的少光下。它提出了一种相关技术的进一步改进，该技术也由Stelzer小组介绍，称为选择性平面照明显微镜(SPIM)(Huisken等人，2004年)。科学，305：1007-9)。新设备将设在基因组学和系统生物学研究所(IGSB)(网址：www.igsb.org/)，并将立即对CCSB的先进成像平台产生影响，原因有三个。首先，它将在很大程度上补充我们现有的共聚焦系统，以及芝加哥大学开发的微流体设备。其次，它将首次使我们能够动态成像模式生物(蠕虫、苍蝇、青蛙和斑马鱼)以及组织和细胞培养中不同生理条件下的荧光标记转录和其他因子，在长时间内具有最高的时空分辨率。最后，DSLM不仅将利用CCSB的研究和基础设施，还将利用NIH资助的其他多个项目，包括本申请中提出的项目，将实时成像显微镜研究提高到下一个水平和分辨率。 与公众健康相关：新开发的荧光数字扫描激光光片显微镜(DSLM)显著减少了对标本的光损害，同时提高了实时成像的速度和质量。这种功能的组合是该仪器独一无二的，它将使芝加哥大学和合作机构的研究人员能够在长时间(例如，几天的开发)和细胞分辨率下进行体内成像实验。这将极大地提高我们在细胞和发育基因组研究中的能力，特别是通过可视化和研究正常和疾病中基因表达的时空网络。

DESCRIPTION (provided by applicant): We request a custom built Digital Scanned Laser Light Sheet Microscope (DSLM), which will enable a broad group of investigators in Chicago to visualize proteins in live tissues and organisms with high resolution and speed and without sample damage associated with other fluorescence microscopy techniques. A major user group will be the investigators participating in at the Chicago Center for Systems Biology (CCSB) (://www.chicago-center-for-systems- biology.org/; grant P50GM081892). The Center's scientific program focuses on the dynamics of transcriptional networks on physiological, developmental and evolutionary scales. The new system will particularly enable us to generate and analyze extensive libraries of the expression networks in development, norm and disease progression. Because of the unique capabilities of this instrument, we have also received requests from groups outside The University of Chicago and we will reserve 25% time on the instrument for these users (minor user group). The DSLM is an innovative live-imaging fluorescence microscopy system developed by Dr. Stelzer's group at the European Molecular Biology Laboratory (EMBL) (Keller et al., 2008. Science, 322:1065-9). In comparison to other advanced fluorescence microscopy techniques (confocal and two-photon) the DSLM provides more than 50 times higher imaging speeds with 10 times higher signal to noise ratio, while exposing the specimens to at least two orders of magnitude less light. It presents a further improvement of a related technology, also introduced by the Stelzer group, known as Selective Plane Illumination Microscopy (SPIM) (Huisken et al., 2004. Science, 305:1007-9). The new equipment will be housed at the Institute for Genomics and Systems Biology (IGSB) (://www.igsb.org/) and will have an immediate impact on CCSB's advanced imaging platform for three important reasons. First, it would largely complement our existing confocal system coupled with a microfluidics device developed at The University of Chicago. Second, it will allow us for the first time to dynamically image fluorescently tagged transcription and other factors in model organisms (worm, fly, frog, and zebrafish) as well as in tissues and cell cultures under different physiological conditions over long periods of time, with the highest spatiotemporal resolution. Finally, the DSLM would leverage not only the CCSB research and infrastructure, but also other multiple NIH funded projects, including those presented in this application, bringing live imaging microscopy studies to the next level of speed and resolution. PUBLIC HEALTH RELEVANCE: The newly developed fluorescence Digital Scanned Laser Light Sheet Microscope (DSLM) dramatically minimizes photo damage to the specimen at the same time increasing both the speed and quality of live imaging. Such combination of features is unique to this instrument, which will enable investigators at The University of Chicago and collaborating institutions to perform in-vivo imaging experiments over long durations (e.g. several days of development)and at cellular resolution. This will greatly enhance our capabilities in cellular and developmental genomic research, particularly through visualizing and studying spatiotemporal networks of gene expression in norm and disease.