The development of hybrid widefield imaging for out-of-focus background rejection

描述（由申请人提供）：摘要本项目的目标是为生物医学界带来一种新的光学成像技术，并将其建立为一种简单，通用和强大的共聚焦显微镜的替代品。我们的技术被称为HiLo成像，是我们实验室在2008年夏天发明的。HiLo成像允许在不使用共焦扫描机制的情况下共焦样离焦荧光背景抑制。简而言之，它需要获取两个图像，一个具有结构化照明，另一个具有标准均匀照明。HiLo成像适用于荧光和非荧光样品。由于它是基于传统的宽视场成像，它可以很容易地适应标准显微镜。此外，由于它只需要两个图像，它可以在非常高的采集速率下工作，很容易超过视频速率（有待证明）。 该项目主要集中在仪器开发上。已经建立了HiLo显微镜的基本原理，我们计划通过实验量化关键性能参数，包括轴向分辨率，信噪比和深度穿透。为了证明HiLo成像的多功能性及其广泛的适用性，我们计划在共聚焦显微镜不切实际或不可能的情况下确认HiLo成像的有效性。具体来说，我们计划建立一个HiLo显微内镜和HiLo宏观镜。前者将用于在大鼠结肠中进行活体共焦成像，与波士顿大学医学院胃肠病学系的Satish Singh博士小组合作，目的是证明HiLo显微内镜用于光学活检和癌症诊断的有效性。我们的第二个设备，一个HiLo宏观，将利用HiLo成像的多功能性，提供什么共聚焦显微镜不能，即视频速率，在一个非常宽的视野（约1厘米）和长的工作距离（几厘米）的焦点外的背景抑制。该宏观镜将应用于小动物的分子成像，我们建议与马萨诸塞州总医院放射肿瘤科的Rakesh Jain博士小组合作，证明其在小鼠肿瘤模型中表征异常血管系统的有效性。 HiLo成像是一项新技术。我们的目标是牢固地确立这项技术的优势，使其成为生物医学成像的常规仪器。 公共卫生相关性：项目叙述公共卫生：我们发明了一种新的光学成像技术，以拒绝失焦的背景，是简单的，廉价的和多功能的。我们建议开发一个内窥镜和一个宽视场的宏观基于此技术的临床前小动物癌症模型成像和诊断。

DESCRIPTION (provided by applicant): ABSTRACT The goal of this project is to bring a new optical imaging technology to the biomedical community, and establish it as a simple, versatile and robust alternative to confocal microscopy. Our technique is called HiLo imaging, and was invented by our lab in the summer of 2008. HiLo imaging allows confocal-like out-of-focus fluorescence background rejection without the use of a confocal scanning mechanism. In brief, it requires the acquisition of two images, one with structured illumination and another with standard uniform illumination. HiLo imaging works with both fluorescent and non-fluorescent samples. Since it is based on conventional widefield imaging, it can readily be adapted to standard microscopes. Moreover, since it requires only two images, it can be operated at very high acquisition rates that easily surpass video rate (to be demonstrated). This project is focused mostly on instrumentation development. Having already established the basic principles of HiLo microscopy, we plan to experimentally quantify key performance parameters, including axial resolution, signal to noise ratio and depth penetration. To demonstrate the versatility of HiLo imaging and its broad range of applicability, we plan to confirm the effectiveness of HiLo imaging in situations where confocal microscopy is impractical or impossible. Specifically, we plan to build both a HiLo endomicroscope and a HiLo macroscope. The former will be used to perform intravital confocal-like imaging in rat colons, in collaboration with the Dr. Satish Singh group at the Boston University School of Medicine Gastroenterology Department, with the goal of demonstrating the effectiveness of HiLo endomicroscopy for optical biopsy and cancer diagnosis. Our second device, a HiLo macroscope, will exploit the versatility of HiLo imaging by providing what confocal microscopy cannot, namely video-rate, out-of-focus background rejection over a very wide field of view (about 1cm) and with a long working distance (several cms). The applications of this macroscope will be in molecular imaging of small animals, and we propose to demonstrate its effectiveness at characterizing abnormal vasculature in mouse tumor models, in collaboration with the Dr. Rakesh Jain group at the Massachusetts General Hospital Department of Radiation Oncology. HiLo imaging is a new technology. Our goal is to firmly establish the advantages of this technology so that it will become routine instrumentation in biomedical imaging. PUBLIC HEALTH RELEVANCE: PROJECT NARRATIVE Public health: We have invented a new optical imaging technology to reject out-of- focus background that is simple, inexpensive and versatile. We propose to develop an endomicroscope and a widefield macroscope based on this technology for pre-clinical small-animal cancer-model imaging and diagnosis.