Enhanced Knife-Edge Scanning Microscopy for Sub-micrometer Imaging of Whole Small Animal Organs

奖项摘要#1256086：用于整个小动物器官亚微米成像的增强刀口扫描显微镜这项NSF奖授予了德克萨斯a&m大学脑网络实验室，将有助于增强和改造刀口扫描显微镜（KESM），一种“独一无二”的显微镜仪器，成为一种更广泛可用的生物研究系统。KESM能够在亚微米分辨率下对整个小动物器官进行切片和成像（也就是说，非常精细，比单个细胞的大小还小）。建立了KESM仪器样机，并成功地在亚微米分辨率下扫描了包括小鼠全脑、章鱼脑和小鼠肺在内的多种生物器官。该项目将大大增强该原型，并将其转变为一个强大的系统，可以被其他研究小组和行业合作伙伴轻松复制和操作。由该项目建立的增强仪器至少解决了生物学研究中的两个主要新兴方向：(1)“组学”（研究某些生物实体的总集合，例如基因基因组学）和(2)多尺度建模（跨一系列尺度的建模系统，例如从细胞到整个器官）。随着基因组学在20世纪90年代及以后取得的巨大成功，生物学研究越来越多地转向各种形式的组学。许多组学研究依赖于解剖学信息（例如，连接组学，研究大脑中的完整接线图）或全器官水平的基因组信息（例如，基因表达水平）。多尺度建模也已成为生物学研究中一个重要的方法论过程。在组学和多尺度建模的许多项目中，整个生物器官的亚微米显微数据是必不可少的，但现有的工具无法满足当前的需求。预计KESM将填补这一空白。该仪器的具体增强包括：(1)增强成像（更高分辨率的光学和相机，荧光成像[遗传和分子研究的关键]），(2)增强机械控制（更严格，更准确，更高分辨率的运动控制），(3)增强切割（振动刀，实时监测），以及(4)增强鲁棒性（质量监测，校准）。总的改进结果可以概括如下：(1)成像分辨率提高了3倍，(2)操作鲁棒性提高了10倍，(3)成像速度提高了10倍（与竞争方法相比），(4)新的荧光成像能力。更广泛的影响：(1)对研究界的影响：增强的KESM将使研究人员能够获得高分辨率的全器官数据，用于不同类型生物器官的多尺度组学研究。(2)对教育的影响：整个生物器官的微观地图集，如项目团队开发的基于网络的KESM小鼠脑地图集，将作为各级学生和教育工作者（K-12，本科生，研究生和公众）的教育资源。作为该项目的一部分，研究生和本科生将在多学科环境（神经科学和计算机科学）中接受培训。(3)仪器推广计划：项目组将与一家初创公司合作，简化KESM仪器的系统集成和制造流程，以实现更广泛的推广。仪器的设计和操作说明将提供给生物研究界的人谁希望建立自己的仪器。

Award Abstract #1256086: Enhanced Knife-Edge Scanning Microscopy for Sub-micrometer Imaging of Whole Small Animal OrgansThis NSF award, made to the Brain Networks Laboratory at Texas A&M University, will help enhance and transform the Knife-Edge Scanning Microscope (KESM), a "one of a kind" microscopy instrument, into a more widely available system for biological research. KESM is capable of sectioning and imaging whole small animal organs at submicrometer resolution (that is, in very fine detail, smaller than the size of a single cell). A prototype KESM instrument was constructed, and its capability was successfully demonstrated by scanning diverse biological organs including whole mouse brains, octopus brains, and the mouse lung at submicrometer resolution. This project will enhance this prototype significantly, and transform it into a robust system that can be replicated and operated with ease by other research groups and industry partners. The enhanced instrument to be built by this project addresses at least two major emerging directions in biological research: (1) "omics" (the study of a total collection of some biological entity, for example, genomics for genes) and (2) multi-scale modeling (modeling systems across a range of scales, for example from the cell up to the whole organ). With the tremendous success of genomics in the 1990's and beyond, biological research is increasingly moving toward various forms of omics. Much of the omics research depends on anatomical information (e.g., connectomics, studying the complete wiring diagram in the brain) or genomic information (e.g., gene expression levels) at the whole-organ-level. Multi-scale modeling has also become a major methodological process in biological research. In many projects in omics and multi-scale modeling, sub-micrometer microscopy data from whole biological organs are essential yet available tools are unable to meet the current demand. KESM is expected to fill this gap. The specific enhancements of the instrument include: (1) enhanced imaging (higher resolution optics and camera, fluorescence imaging [key to genetic and molecular studies]), (2) enhanced mechanical control (more rigid, accurate, and higher resolution motion control), (3) enhanced cutting (vibrating knife, real time monitoring), and (4) enhanced robustness (quality monitoring, calibration). The overall resulting improvement can be summarized as follows: (1) 3X improvement in imaging resolution, (2) 10X improvement in robustness of operation, (3) 10X improvement in imaging speed (compared to competing methods), (4) new fluorescence imaging capability. Broader impacts: (1) Impact on the research community: The enhanced KESM will allow researchers to obtain high resolution, whole-organ data for multi-scale, omics investigation of various types of biological organs. (2) Impact on education: Microscopic atlases of whole biological organs, such as the web-based KESM mouse brain atlas developed by the project team, will serve as an educational resource for students and educators at all levels (K-12, undergraduate, graduate, and general public). As part of this project, graduate and undergraduate students will be trained in a multidisciplinary environment (neuroscience and computer science). (3) Instrument dissemination plan: The project team will collaborate with a start-up company, to streamline system integration and manufacturing of the KESM instrument for broader dissemination. The design of the instrument and the operational instructions will be made available to the biological research community for those who wish to build their own instrument.