Live Imaging Core

项目摘要/摘要(实时成像核心) 实时成像核心提供有关使用、监督访问和维护最新状态的培训 艺术成像设备，以及仪器和分析方法的不断发展。这个 这个核心的三个主要设备组件，定制的薄片荧光显微镜， 用于光学薄片显微镜的高通量样品处理的仪器，以及用于低通量样品处理的工具。 放大荧光成像是实现研究项目目标的核心。技术上的 该核心的专业知识将确保所有项目研究都是在推动 斑马鱼肠道内微生物可视化的当前边界。我们已经演示过了 以单细菌分辨率显示斑马鱼幼体肠道微生物的能力，观察细菌 增长和竞争，并从结果图像中提取定量信息；这些功能 是我们提议的项目实验所必需的。因为我们的仪器是定制设计和制造的， 它为实验者提供了最大的灵活性，包括使用多个荧光探针，以及集成 定制编写的软件来控制各种硬件，以适合所提议的实验的方式。在……里面 此外，图像分析代码也是通过定制开发的软件来实现的，该软件利用半 自动化方法和基于机器学习的分类对象；正在进行的工作旨在进一步优化 这些方法。我们的客户开发的高通量方法解决了成像大容量的挑战 我们正在开发低倍率的“宏观”方法，它使用简单的 然而，多用途的相机和过滤器，以快速评估荧光生物的丰富性在广泛的领域 查看。这一核心将继续设计和改进仪器。同时，核心人员将进行培训 安排项目研究人员使用该仪器，并与他们一起设计实验 最大限度地提高效率和生产力。实现这一核心的目标将显著增强 能够进行研究项目中描述的实验。

PROJECT SUMMARY/ABSTRACT (LIVE IMAGING CORE) The Live Imaging Core provides training in the use of, supervised access to, and maintenance of state-of-the- art imaging equipment, as well as the continued development of instrumentation and analysis methods. The three major equipment components of this Core, custom-built light sheet fluorescence microscopes, instrumentation for high throughput sample handling for light sheet microscopy, and tools for low- magnification fluorescence imaging are central to achieving the aims of the Research Projects. The technical expertise of this Core will ensure that all Project research is carried out on instrumentation that pushes the current boundaries for visualization of microbes within the zebrafish intestine. We have already demonstrated the ability to visualize larval zebrafish intestinal microbes with single-bacterium resolution, watching bacterial growth and competition and extracting quantitative information from the resulting images; these capabilities are requisite for our proposed Project experiments. Because our instrumentation is custom designed and built, it allows maximum flexibility for experimenters, including use of multiple fluorescent probes, and integration of custom-written software to control various hardware in ways tailored to the proposed experiments. In addition, image analysis code is also achieved by custom-developed software that makes use of semi- automated methods and machine-learning-based classification objects; ongoing work aims to further optimize these approaches. Our custom developed high throughput methods address the challenge of imaging large numbers of specimens and we are developing low-magnification “macroscope” methods that employ simple yet versatile cameras and filters to rapidly assess abundances of fluorescent organisms over wide fields of view. This Core will continue to design and refine instrumentation. At the same time Core personnel will train Program Project research personnel in use of this instrumentation, and work with them to design experiments that maximize efficiency and productivity. Achieving the aims of this Core will significantly enhance the ability to carry out experiments described in the Research Projects.