Tissue protein imaging via second harmonic generaton

描述(由申请人提供)： 组织蛋白质结构，如胶原和肌动球蛋白，产生强烈的二次谐波(SHG)，由此产生的对比度在激光扫描SHG显微镜上产生高分辨率的活组织图像。描述倍频的物理学表明，关于蛋白质二级和高阶结构的信息可以从这些蛋白质与特定的光偏振相互作用中推导出来。获取激光扫描图像的小型化技术正在迅速发展。因此，如果有必要，一项专门的工程工作可能很快就会允许医生分析活着的患者肌肉和结缔组织中的微观和亚微观结构。然而，现在的关键问题是：i)是否可以从SHG图像中获得有医学价值的信息；ii)哪些组织和医疗条件最容易分析；iii)收集每个组织中的SHG图像的最优和最简单的方法；iv)SHG的哪些物理性质可能导致对天然蛋白质结构的重要医学见解；以及v)精细结构分析所需的光学是否适用于活组织。这些问题定义了两条基本路径。首先是不同组织(包括发育状态和疾病状态)的倍频图像特征，以及从每种蛋白质/组织类型生成倍频对比图像的参数的优化。其次是从组织蛋白质阵列中分析倍频的分子和物理基础，并检验倍频偏振分析对探测组织中蛋白质组件的亚微观结构的预测能力。我们的具体目标遵循每一条有希望的道路，并将寻求两者之间的趋同。这些目的是i)测试SHG成像作为一种诊断手段的有效性，以揭示特定组织的组织学/病理学：骨骼和心肌、皮肤、软骨、骨骼和疾病引起的斑块沉积。Ii)使用详细的偏振分析和超分辨技术来表征组织中倍频的分子来源、构成这种光学活性的这些分子的结构状态，以及它们的光学性质对分子结构变化的响应。Iii)确定活体组织倍频成像的理想光学参数。

DESCRIPTION (provided by applicant): Tissue protein structures such as collagen and actomyosin give rise to intense second harmonic generation (SHG), and the resulting contrast yields high-resolution images of live tissues on a laser-scanning SHG microscope. The physics describing SHG indicates that information on secondary and higher-order protein structure may be deduced from the interactions of these proteins with specific polarizations of light. Miniaturized technology to acquire laser-scanning images is developing quickly. Therefore, if warranted, a dedicated engineering effort could soon permit physicians to analyze microscopic and submicroscopic structure in muscle and connective tissues of live patients. However, the key questions now are: i) whether medically valuable information can be derived from SHG images, ii) which tissues and medical conditions are most readily analyzed, iii) what the most optimal and simplest means will be for collecting SHG images in each tissue, iv) which of the physical properties of SHG might lead to medically-important insights into native protein structure, and v) whether the optics required for fine structural analysis will apply in the context of live tissues. These questions define two basic paths. First is characterization of SHG image profiles from various tissues (including developmental and diseased states), and optimization of parameters for generating SHG contrast images from each protein/tissue type. Second is analysis of the molecular and physical basis of SHG from tissue protein arrays, and testing of the predicted power of SHG polarization analysis to probe sub-microscopic structure of protein assemblies in tissues. Our specific aims follow each of these promising paths, and will seek convergence between the two. These aims are i) Test the utility of SHG imaging as a diagnostic modality to reveal histology/pathology of specific tissues: skeletal and cardiac muscle, skin, cartilage, bone, and plaque deposits caused by disease. ii) Use detailed polarization analysis and super-resolution techniques to characterize the molecular sources of SHG in tissues, the structural states of these molecules that underlie this optical activity, and the response of their optical properties to changes in molecular structure. iii) Determine ideal optical parameters for SHG imaging of tissues in vivo.