Towards Deep Tissue Imaging of Fluorescence Resonance Energy Transfer Parameters

[0854249webb]荧光共振能量转移（FRET）已被证明在研究化学物质向细胞的转运以及诸如蛋白质错误折叠（可能导致阿尔茨海默病）等疾病的潜在原因方面具有巨大价值。已经测量了FRET转移对供体荧光的猝灭，分子内FRET参数（寿命和产率，在柔性连接体的情况下，连接体距离的均值和方差）已被广泛使用。然而，FRET成像已被限制在体外或表面显微镜，因为实质性的散射有害的影响。该项目的目标是展示一种在高散射介质中成像FRET参数的方法，从而为深部组织体内FRET研究提供机会。新的工作计划如下：-开发一种用ODT成像FRET参数的方法：将开发一种荧光共振能量转移光学扩散层析成像（FRET-ODT）成像方法。这将涉及到分子内FRET参数的解决方案，包括刚性和柔性连接体，这些连接体作为未知源纳入供体荧光的扩散方程表示中。将研究对参数空间（调制频率、源-检测器排列、FRET参数、信噪比）的灵敏度，并开发适用于散射介质中FRET源实验数据的软件。结合受体动力学的优点也将被研究。以类似的方式，分子间FRET源将在ODT框架中实现。- FRET-ODT实验研究：将构建一个模型组织实验，以获得数据来评估将要开发的FRET-ODT成像方法。将获得允许不同程度的散射、不同的FRET化学浓度、不同的供体-受体距离和距离分布、多个调制频率以及非均匀背景的影响的数据。作为体内FRET- odt应用，将使用FRET模型研究靶向药物递送到癌细胞的小鼠模型，以及随后的药物进入和释放，该模型包含有关运输动力学的信息，并模拟药物递送过程。这项研究的信息将有助于开发靶向药物，选择性地将其传递给癌细胞，从而防止化疗期间通常发生的对健康细胞的损害（例如，白细胞减少-白细胞缺乏，脱发-脱发，便秘和高血压）。这项工作将由普渡大学化学系的Low小组完成。

0854249WebbFluorescence resonance energy transfer (FRET) has proved to be of immense value in the study of chemical transport into cells and the underlying cause of disease due to, for example, protein misfolding (which may lead to Alzheimer's disease). The quenching of donor fluorescence by FRET transfer has been measured, and the intramolecular FRET parameters (lifetime and yield, and in the case of a flexible linker, the mean and variance of the linker distance) have been widely used. However, FRET imaging has been limited to in vitro or surface microscopy because of the deleterious effects of substantial scatter. The goal of this project is to demonstrate a method for imaging FRET parameters in highly scattering media, thereby providing the opportunity for deep tissue in vivo FRET studies. The new work planned is as follows:- Development on an Approach to Image FRET Parameters with ODT: A fluorescence resonance energy transfer optical diffusion tomography (FRET-ODT) imaging approach will be developed. This will involve a solution for the intramolecular FRET parameters with both rigid and flexible linkers that are incorporated as unknown sources in a diffusion equation representation for the donor fluorescence. The sensitivity to the parameter space (modulation frequencies, source-detector arrangement, FRET parameters, signal to noise ratio) will be studied and software suitable for use with experimental data for FRET sources in a scattering medium will be developed. The merits of incorporating acceptor kinetics will also be investigated. In a similar way, intermolecular FRET sources will be implemented in an ODT framework.- FRET-ODT Experimental Studies: A model tissue experiment will be constructed to obtain data to evaluate the FRET-ODT imaging approach to be developed. Data allowing the influence of varying degrees of scatter, varying FRET chemical concentration, varying donor-acceptor distance and distance distribution, and multiple modulation frequencies, as well as the effect of an inhomogeneous background, will be obtained. As an in vivo FRET-ODT application, a mouse model for targeted drug delivery to cancer cells, with the subsequent drug entry and release, will be studied using a FRET model that incorporates information about the transport kinetics and which emulates the drug delivery process. Information from this study will aid in the development of targeted drugs that are selectively delivered to cancer cells, thereby preventing damage to healthy cells that usually occurs during chemotherapy (for example, leucopenia - a deficiency in white blood cells, alopecia - hair loss, constipation, and hypertension). This work will be done with the Low group in the Dept. of Chemistry at Purdue.