Time-gated confocal Raman microscope

描述（由申请人提供）：生命科学的进展取决于新工具和仪器的开发。通过能够提供体内结构和化学信息的成像技术，我们在细胞和分子水平上理解生命系统功能的能力大大增强。拉曼光谱是真正的非侵入性的，它可以提供重要的信息的化学组成和物理结构的生物组织。这项小规模（R03）研究提案旨在开发一种非侵入性显微光谱学的创新方法，以明确探索细胞和组织中化学成分的变化。为了实现这一目标，PI将利用时间选通拉曼成像的概念，首次应用于显微镜，以实现对荧光背景具有上级分辨力的共焦成像。预期它将提供信噪比的至少一个数量级的改进，这转换成至少快100倍的采集速率，同时提高空间分辨率并消除在解析复杂拉曼频带中可能的模糊性。第一个具体目标是建立一个新的仪器的原型和表征其性能。第二个具体目标是对用于几种模型系统的分子、细胞和组织成像的新仪器进行试点测试：1）胶原蛋白自组织和结构转化，这导致骨结构发育，2）视网膜色素上皮细胞的光降解，这被认为是失明的主要原因，以及3）牙齿组织中的早期龋齿检测。许多医学研究和诊断应用需要具有最小侵入性的显微分子成像。拉曼显微光谱是一种非侵入性的、化学特异性的分子成像技术，但由于强荧光背景而存在显著的缺点，这显著降低了信噪比，并且难以揭示振动带的精细结构。拟议的研究提供了一个解决这个问题的显微成像的时间门控有用的信号在一个专门设计的光学装置。

DESCRIPTION (provided by applicant): Progress in the Life Sciences depends upon the development of new tools and instruments. Our ability to understand the function of living systems on a cellular and molecular level is greatly enhanced by imaging techniques capable of providing structural and chemical information in vivo. Raman spectroscopy is truly non-invasive, and it could provide significant information on the chemical composition and physical structure of biological tissues. This small-scale (R03) research proposal aims to develop an innovative approach for non-invasive microspectroscopy in order to unambiguously explore changes in chemical composition in cells and tissues. To achieve this goal, the PI will utilize the concept of time-gated Raman imaging, applied for the first time to microscopy to achieve confocal imaging with superior discrimination against the fluorescence background. It is expected that it will provide at least an order of magnitude improvement in the signal-to-noise ratio, which transforms into at least 100 times faster acquisition rates, while improving spatial resolution and eradicating possible ambiguities in resolving complex Raman bands. The first specific aim is building the prototype of a novel instrument and characterizing its performance. The second specific aim is pilot testing of a new instrument for molecular, cellular and tissue imaging for several model systems: 1) collagen self-organization and structural transformation, which leads to bone structure development, 2) photodegradation of retinal pigment epithelium cells, which is considered to be the major cause of blindness, and 3) early caries detection in dental tissues. Many medical research and diagnostic applications require microscopic molecular imaging with minimum invasiveness. Raman microspectroscopy, being a non-invasive, chemically specific technique for molecular imaging, suffers a dramatic drawback due to a strong fluorescent background, which significantly reduces the signal-to-noise ratio and makes it difficult to reveal the fine structure of vibrational bands. The proposed research provides a solution to this problem for microscopic imaging by time-gating the useful signal in a specially designed optical arrangement.