Targeted Multi-Spectral Dual Axes Confocal Imaging of In Vivo Molecular Expressio

描述（由申请人提供）：体内分子表达的靶向多光谱双轴共聚焦成像本BRP的广泛、长期目标是开发一种微型活体成像仪器，以研究活体动物上皮中癌症生物学的分子机制。这种新颖的光学设计使用离轴照明和光的收集来产生用于收集垂直横截面荧光图像的上级动态范围。我们将组建一个由密歇根大学工程学院和医学院的研究人员组成的跨学科团队。具体目的是1）开发扫描机制以实现真实的时间、多光谱垂直横截面成像，2）选择与结肠粘膜上皮中的异型增生和乳腺血管内皮中的CXCR 7结合的亲和肽，和3）验证活体动物模型中的成像性能。双轴共焦结构使用两个独立的低数值孔径物镜的重叠焦点来实现亚细胞分辨率和长工作距离。后物镜扫描提供了一个大的视场以及光学器件的可扩展性和小型化，并且复制的抛物面镜将准直的多光谱光束引导到一个共同的焦点。这种独特的设计可以观察基底到管腔方向的上皮分化模式。上皮的真实的时间成像将通过开发垂直致动器来实现，该垂直致动器使用具有克服运动伪影（>10帧/秒）的尺寸、速度、力和线性的薄膜压电材料。该器件将与由参数谐振驱动的横向扫描微镜相结合。亲和肽将使用噬菌体展示技术，通过针对过表达与瘤形成相关的表面靶标的细胞和组织生物淘选高多样性文库来选择。多个探针可以被近红外标记以实现预期的组织穿透深度（>500微米）。将通过将微型原型插入结肠以评价远端粘膜上皮或通过贴靠胸部放置以对活体动物乳腺导管上皮成像来验证成像性能。公共卫生：这种通用仪器可用于研究活体动物模型上皮细胞中的分子过程，具有亚细胞分辨率，高动态范围和全成像深度，允许纵向研究癌症生物学机制，并为药物发现提供新平台。 公共卫生相关性： 这些目标的完成将为活体动物上皮成像提供一种具有上级深度和动态范围的通用仪器，从而可以研究癌症的分子机制，并为药物发现提供一个新的平台。

DESCRIPTION (provided by applicant): Targeted Multi-Spectral Dual Axes Confocal Imaging of In Vivo Molecular Expression The broad, long-term objective of this BRP is to develop a miniature intra-vital imaging instrument to study molecular mechanisms of cancer biology in the epithelium of live animals. This novel optical design uses off-axis illumination and collection of light to produce superior dynamic range for collecting vertical cross-sectional fluorescence images. We will assemble an inter-disciplinary team of investigators from the College of Engineering and the School of Medicine at the University of Michigan. The specific aims are 1) to develop a scanning mechanism to achieve real time, multi- spectral vertical cross-sectional imaging, 2) to select affinity peptides that bind to dysplasia in colonic mucosal epithelium and to CXCR7 in breast vascular endothelium, and 3) to validate imaging performance in live animal models. The dual axes confocal architecture uses the overlapping focus of two separate low numerical aperture objectives to achieve sub-cellular resolution and long working distance. Post-objective scanning provides a large field-of-view as well as scalability and miniaturization of the optics, and a replicated parabolic mirror directs collimated, multi-spectral beams to a common focus. This unique design can view the epithelial differentiation pattern in the basilar to luminal direction. Real time imaging of the epithelium will be achieved by developing a vertical actuator that uses thin film piezoelectric materials that have the size, speed, force, and linearity to overcome motion artifact (>10 frames/sec). This device will be combined with a lateral scanning micro-mirror that is driven by parametric resonance. Affinity peptides will be selected using the technique of phage display by biopanning a high diversity library against cells and tissues that over express surface targets associated with neoplasia. Multiple probes can be near-infrared labeled to achieve the tissue penetration depths expected (>500 microns). Imaging performance will be validated by inserting the miniature prototype into the colon to evaluate the epithelium of the distal mucosa or by placing against the chest to image the epithelium of breast ducts of live animals. Public Health: This general purpose instrument can be used to study molecular processes in the epithelium of live animal models with sub-cellular resolution, high dynamic range, and full imaging depth, allowing for longitudinal investigation of mechanisms of cancer biology and providing a new platform for drug discovery. PUBLIC HEALTH RELEVANCE: Project Narrative Completion of these aims will deliver a general purpose instrument for imaging the epithelium in live animals with superior depth and dynamic range, allowing for study of molecular mechanisms of cancer and providing a new platform for drug discovery.