In Vivo Immunofluorescence Microscopy and Cytometry

描述（由申请人提供）： 该多学科研究计划的总体长期目标是应用新的光学技术来研究与细胞免疫反应相关的早期变化，并在体内和细胞水平上监测随时间推移对治疗的反应。具体而言，我们提出了在体内免疫荧光显微镜成像血管和淋巴管内皮细胞，循环白细胞和组织树突状细胞表达的特定细胞表面标志物。能够实现非侵入性实时细胞成像的技术是我们实验室最近开发的视频速率共聚焦和多光子荧光显微镜。我们将应用这项技术来研究体内细胞免疫应答调节的重要步骤，包括内皮细胞活化，白细胞-内皮细胞相互作用，树突状细胞募集和迁移。此外，我们建议开发一种体内流式细胞仪，一种新的技术，用于实时，非侵入性检测和定量的循环细胞标记的荧光抗体或抗体片段。我们将使用该系统来监测特定T细胞群体响应免疫调节的变化，检测循环肿瘤细胞，并研究肿瘤细胞脱落和转移潜力之间的相关性。

DESCRIPTION (provided by applicant): The overall long-term goal of this multidisciplinary research program is to apply new optical technology to investigate early changes associated with cellular immune response and to monitor response to therapy over time in vivo and at the cellular level. Specifically, we proposed to develop in vivo immunofluorescence microscopy for imaging specific cell surface markers expressed by vascular and lymphatic endothelial cells, circulating leukocytes, and tissue dendritic cells. The technology that enables noninvasive, real-time cellular imaging is a video rate confocal and multiphoton fluorescence microscope that we recently developed in our laboratory. We will apply this technology to study important steps involved in the regulation of cellular immune response in vivo, including endothelial cell activation, leukocyteendothelial interaction, and dendritic cell recruitment and migration. In addition, we proposed to develop an in vivo flow cytometer, a novel technology for real-time, noninvasive detection and quantification of circulating cells that are tagged with fluorescent antibodies or antibody fragments. We will use this system to monitor changes in specific T cell population in response to immunomodulation, to detect circulating tumor cells and to investigate the correlation between tumor cell shedding and metastatic potential.