OPTICAL SECTIONING THAT PRESERVES USEFUL DEPTH OF FIELD

(该应用程序是对NIMH项目公告PA-99-007的响应，该项目名为“用于大脑显微成像的探头和仪器”。)我们设计了一种基于广域光学原理和LED照明的三维显微样品光学切片的新策略。高对比度标本的整体结构(例如带有大脑切片的染色神经元)将被成像为在彩色(蓝色)背景下显示的黑色轮廓。焦平面内的结构将以相反的颜色(黄色)成像，并通过黑暗的轮廓显示为薄的光学部分。光场内的结构将以高分辨率成像，而截面外的结构将以大景深成像。这种方法将在许多情况下补充共聚焦显微镜。通过提供有用的景深信息，我们的方法将允许在整个三维结构的背景下解释每个光学部分。使用广域采光方法，可以实时完成图像采集，图像不需要进一步的数字处理。此外，由于可以通过对现有显微镜进行相对廉价的改进来实现实施，因此可以使更多的科学家更容易获得显微镜标本的高分辨率光学切片。拟议的商业应用：能够以高分辨率实时光学切割三维结构，同时保持有用的景深，这将使使用光学显微镜的生物医学界的大部分人感兴趣。这样的选择很可能会被纳入S的大量新显微镜中，这是一项每年5亿美元的业务。一个更大的市场可能是对现有显微镜的改装。

(This application is in response to NIMH program announcement PA-99- 007, "Probes and instruments for micro-imaging the brain.") We have devised a novel strategy for optical sectioning of three- dimensional microscopic specimens that is based on wide-filed optical principles and LED illumination. The overall structure of a high-contrast specimens (such as a stained neuron with a brain slice) will be imaged as a dark silhouette displayed against a colored (blue) background. Structures within the plane of focus will be imaged in an opposing color (yellow) and displayed as a thin optical section through the dark silhouette. The structures within the optical field will be imaged at high resolution, whereas structures outside the section will be imaged with great depth of field. This method will complement confocal microscopy in many situations. By providing useful depth-of-field information, our approach will allow each optical section to be interpreted in the context of the overall three-dimensional structure. With the use of the wide-field approach for light gathering, image acquisition can be completed in real time and images will not require further digital processing. Moreover, because implementation can be achieved with relatively inexpensive modifications to existing microscopes, high-resolution optical sectioning of microscopic specimens an be made more accessible to a greater number of scientists. PROPOSED COMMERCIAL APPLICATIONS: The ability to optically section a three-dimensional structure with high resolution in real time while preserving useful depth-of-field would interest a major portion of the biomedical community who use optical microscopes. Such an option would likely be incorporated in s significant number of new microscopes with is a $500 million a year business. An even larger market may be the retrofitting of existing microscopes.