DEVELOPMENT OF A HOLOGRAPHIC FLUORESCENCE MICROSCOPE

这个奖支持开发一种新型荧光显微镜。这种被称为Holoskop的显微镜，一旦开发出来，将允许在三维（3D）中实时观察微观标本。在过去的十年里，细胞科学的进步已经清楚地表明，小分子和蛋白质在细胞内的快速运动。然而，目前还没有显微镜可以实时观察这些事件。从水母和其他水生生物（GFP）中获得的绿色荧光蛋白已被分离和测序。因此，通过将来自GFP的DNA与感兴趣的蛋白质的DNA连接起来，GFP可以作为生物感兴趣的蛋白质表达的一部分在细胞中表达。这种表达使人们能够通过跟踪GFP标签的荧光来跟踪具有生物学重要性的靶蛋白的运动。这些荧光蛋白构建探针和其他有机荧光探针，结合利用另一种称为共聚焦显微镜的光学切片技术的进步，已经彻底改变了蛋白质、离子和信使分子的3D定位和动态运输研究。在实践中，探针的三维成像来源于通过z平面在细胞或组织中拍摄的图像堆栈。因此，识别细胞、结构或过程的荧光分子，在图像堆栈捕获期间在x-y或x-y-z平面上快速移动，目前还不能充分解决或研究。了解这些快速过程及其相互作用对于理解细胞和组织正常和异常功能中的无数事件至关重要。该项目的目标是开发一种新的、明显更快的3D成像显微镜。Holoskop将利用扫描全息原理加速3D成像，而不需要收集图像堆栈。除了开发一种重要的生物学新工具外，该项目还将对参与该项目的大学固有的教育和推广活动产生相当大的影响。Holoskop一旦开发出来，将有助于各种生物科学研究中的数十个项目，因为目前还没有方法可以从该仪器中获得期望的那种信息。

This award supports the development of a new kind of fluorescent microscope. This microscope, dubbed the Holoskop, when developed, will allow the viewing of microscopic specimens in 3 dimensions (3D) in real-time.In the last decade, advances in cell science have made it apparent that there is rapid intracellular movement of small molecules and proteins. However there is currently no microscope available to view these events in real time. Green fluorescent proteins obtained from jellyfish and other aquatic organisms (GFP) have been isolated and sequenced. Thus GFP can be expressed in cells as part of the expression of proteins of biological interest by linking the DNA from GFP with the DNA of the protein of interest. Such expression enables one to follow the movement of target proteins of biological importance by following the fluorescence of the GFP tag.These fluorescent protein construct probes, and other organic fluorescent probes, combined with advances in optical sectioning utilizing another type of microscopy called confocal microscopy, have revolutionized studies in 3D localization and dynamic trafficking of proteins, ions, and messenger molecules. In practice, the 3D imaging of probes is derived from stacks of images taken through the z-plane in cells or tissues. Thus fluorescent molecules identifying cells, structures or processes, which move rapidly in the x-y or x-y-z plane during the capture of the image stack, cannot at present be adequately resolved or studied. Knowledge of these fast processes and their interactions is of paramount importance to understanding the myriad of events in the normal and abnormal functioning of cells and tissues. The objective of this project is to develop a new and demonstrably faster microscope for 3D imaging. The Holoskop will utilize scanning holographic principles for accelerated 3D imaging without the need for collection of image stacks.In addition to developing an important new tool for biology, this project will have considerable impact upon the educational and outreach activities inherent in the universities involved in this project. The Holoskop once developed, will aid scores of projects in a wide variety of biological science investigations since there is currently no method that can obtain the kind of information expected from this instrument.