High-resolution spectroscopic imaging with infrared nonlinear optical (IR-NLO) microscopy

项目摘要 作为一种真正的无标记成像技术，振动显微镜提供了细胞和组织的图谱 具有极高的化学对比度，因为它直接探测了 样本。振动成像方法包括红外吸收显微光谱分析和共聚焦 拉曼显微镜，已经成功商业化的方法(正在增长的5亿 美元市场)，现在是分析和生物实验室中常见的调查工具。 在过去的四十年里，这些技术在生物学领域产生了可衡量的影响 和生物医学，提供健康和患病组织的空间分辨率评估 分子视角。这项提议旨在显著提高振动台的能力 显微镜。我们提出了一种新的成像方法，它融合了IR的理想特性 吸收显微镜具有相干、非线性光学(NLO)的一些独特性质 分子的激发。这种新的IR-NLO技术提高了红外吸收的空间分辨率 显微技术提高了十倍，同时对指纹分子振动的敏感度比 基于拉曼的显微方法。 我们的团队由相干拉曼散射显微镜和红外专家组成 显微光谱成像。我们的创新使快速获取红外吸收图像成为可能 分辨率为0.5微米或更高的指纹振动模式。初步数据 结果表明，红外非线性光学方法可以成功地应用于快速激光扫描显微镜， 从而可以方便地对组织样本进行振动成像。在我们的提案中，我们开发、测试和 改进IR-NLO新技术。该技术的验证是通过广泛的 生物医学成像研究，并与当今最先进的红外显微镜进行比较。 拟议的计划解决了红外光谱显微技术中的一个主要挑战，即 提高成像分辨率。这一新功能非常重要，因为更高的分辨率使 鉴定组织中亚微米级的细胞内和细胞外结构，到目前为止 在红外成像中仍然是隐形的。因此，高分辨率成像特性显著 提高了该技术的诊断能力。通过设置新的分辨率标准 指纹振动成像，IR-NLO技术很可能对组织产生重大影响 成像，并可使其在病理学的研究和临床领域使用。

Project Summary A genuine label-free imaging technology, vibrational microscopy provides maps of cells and tissues with exceptionally high chemical contrast as it directly probes the fundamental vibrational modes of samples. Vibrational imaging approaches include IR-absorption micro-spectroscopy and confocal Raman microscopy, methods that have been successfully commercialized (a growing 500 million dollar market) and are now common tools of inquiry found in analytical and biological laboratories. Over the past four decades, these techniques have had a measurable impact in the fields of biology and biomedicine, offering a spatially resolved assessment of healthy and diseased tissues from a molecular perspective. This proposal aims to significantly improve the capabilities of vibrational microscopy. We propose a new imaging approach that merges the desirable properties of IR absorption microscopy with some of the unique properties of coherent, nonlinear optical (NLO) excitation of molecules. This novel IR-NLO technique improves the spatial resolution of IR absorption microscopy by tenfold, while offering higher sensitivity to fingerprint molecular vibrations relative to Raman-based microscopy methods. Our team is comprised of experts in coherent Raman scattering microscopy and IR microspectroscopic imaging. Our innovation makes it possible to rapidly acquire IR absorption images of fingerprint vibrational modes with a resolution of 0.5 micrometer or better. The preliminary data shows that the IR-NLO approach can be successfully adopted in a rapid laser-scanning microscope, allowing convenient vibrational imaging of tissue specimens. In our proposal we develop, test, and improve the new IR-NLO technology. The validation of the technology is achieved through extensive biomedical imaging studies and comparison with the state of the art IR microscopy available today. The proposed program tackles a major challenge in IR spectroscopic microscopy, namely the improvement of imaging resolution. This new capability is significant, as the higher resolution enables the identification of sub-micrometer intra- and extra-cellular structures in the tissue, which hitherto have remained invisible in IR-imaging. The high-resolution imaging property thus dramatically improves the diagnostic capabilities of the technique. By setting a new resolution standard for fingerprint vibrational imaging, the IR-NLO technology is likely to have a significant impact in tissue imaging and can enable its use in both research and clinical domains for pathology.