Compact all-fiber based CARS microscope for biomedical imaging

描述(申请人提供)：荧光成像是目前细胞和组织分析的主要光学方法。它依赖于荧光分子探针的使用，这些荧光分子探针要么由经过修饰的基因表达，要么被标记到感兴趣的分子上。这种修改可能会干扰正常功能，并阻碍对实验结果的解释。此外，荧光团本身会受到光漂白的影响，这会导致成像时间有限，并阻碍长期研究。相干反斯托克斯拉曼散射(CARS)显微镜是一种非线性光学成像技术，可以对分子进行高速振动成像。CARS显微镜有几个独特的优点：(A)CARS成像是免标记和非破坏性的。(B)对激发强度的非线性依赖确保CARS信号仅在焦点中心产生，从而提供固有的亚微米3D空间分辨率。(C)CARS成像提供固有的化学选择性，并允许特定于键的对比度。因此，CARS成像有可能成为一种对细胞和组织进行微创和定量监测的重要方法，这对生物学研究和医学诊断至关重要。这项技术被生物医学研究人员广泛采用的关键障碍是现有激光系统在产生所需的可调谐、高功率和短脉冲激光方面的技术和操作限制。这些系统体积庞大，价格昂贵，需要定期维护。领先的便携式拉曼仪器制造商安捷伦与普渡大学合作，计划通过开发紧凑、坚固且经济实惠的基于光纤激光的CARS显微镜，将CARS研究级实验室仪器推向市场。在第一阶段，我们将通过开发和制作几个目前还没有商业化的光纤组件的原型，然后构建一个功能性的全光纤汽车系统，来证明这一努力的可行性。我们将演示细胞和组织的化学选择性、高对比度和空间分辨率成像。在第二阶段，我们将致力于设计优化，以极低的成本制造相关部件，并提高性能，目标是将这项强大的技术商业化。我们还将设计和展示一种微型光纤CARS成像内窥镜，以便于在临床环境中进行微创原位诊断。拟议研究的成功实施将带来坚固可靠的化学成像显微镜和内窥镜，这些显微镜和内窥镜提供关于体内和体外细胞和 纸巾。 与公共健康相关：对细胞和组织进行非侵入性、实时、成分和亚细胞测量的能力将极大地促进我们对肥胖和相关疾病的了解，提供对动脉粥样硬化的快速准确诊断，并增强对乳房、肺、结肠和喉部肿瘤的检测和诊断。全纤维显微镜还将允许对制药、环境和其他研究领域的样品进行高速光谱分析和化学成像。

DESCRIPTION (provided by applicant): Fluorescence imaging is the current major optical method for cell and tissue analysis. It relies on the use of fluorescent molecular probes which are either expressed by genes which have been modified, or are tagged on to molecules of interest. The modification can interfere with normal function and hamper the interpretation of experimental results. Furthermore, the fluorophores themselves are subject to photobleaching which results in limited imaging time and prevents long term studies. Coherent anti-Stokes Raman scattering (CARS) microscopy is a nonlinear optical imaging technique that allows high-speed vibrational imaging of molecules. CARS microscopy offers several unique advantages: (a) CARS imaging is label-free and non-destructive. (b) The nonlinear dependence on excitation intensity ensures that the CARS signal is only generated in the focal center, providing an inherent sub-micron 3D spatial resolution. (c) CARS imaging offers intrinsic chemical selectivity and allows bond-specific contrast. Thus CARS imaging has the potential to become a vital method for minimal invasive and quantitative monitoring of cell and tissue which is critical for biological research and medical diagnostics. The key hurdles for this enabling technology to be widely adopted by biomedical researchers are the technical and operational limitations of the existing laser systems to generate the required tunable, high-power, and short-pulsed laser beams. These systems are bulky, expensive and require regular maintenance. Agiltron, a leading portable Raman instrument producer, in collaboration with Purdue University, proposes to bring the CARS research level laboratory instrument into the market by developing a compact, robust and affordable fiber laser based CARS microscope. In Phase I we will demonstrate the feasibility of this effort by developing and prototyping several fiberoptic components that are currently not commercially available and then constructing a functional all fiberoptic CARS system. We will demonstrate chemical selective, high contrast and spatially resolution imaging of cells and tissues. In Phase II, we will address design optimization, fabrication of associated components at drastically lower cost, and performance improvement with the goal of commercializing this powerful technique. We will also design and demonstrate a miniaturized fiber CARS imaging endoscope to facilitate minimal invasive in-situ diagnostics in clinical environment. The successful implementation of the proposed research will result in robust and reliable chemical imaging microscopes and endoscopes that provide real time, quantitative and localized structural and molecular information about in-vivo and in-vitro cell and tissues. PUBLIC HEALTH RELEVANCE: The capability to perform non-invasive, real-time, compositional, and sub-cellular measurement of cell and tissue will greatly facilitate our understanding of obesity and related diseases, provide rapid and accurate diagnostics of atherosclerosis, and enhance the detection and diagnosis of tumor in breast, lung, colon, and throat .The all fiber microscope will also allow high-speed spectral analysis and chemical imaging of samples in pharmaceutical, environmental, and other research fields.