Spectroscopic Quantification of Ligand Binding in Vivo

项目总结(见说明)； 该项目的目的是开发新的方法来定量成像靶向磁性纳米颗粒 (MNP)在体内的分布，以提供对何处的机械性洞察 有针对性的MNPs收集以及如何改进有针对性的交付。该项目将结合两部小说 协同方法、光学比率荧光光谱和磁光谱 MNP布朗运动(MSB)，以量化总体浓度吸收，然后量化特定的水平 在体内与靶配体结合，最初专注于可以内化的HER2/neu靶向mNPs 当被捆绑时。这里提出的两种测量技术都是在达特茅斯大学和 我们有相当多的现有资源和专业知识可以利用。 全局分布和结合分数将用OFS通过注入非靶向和 用不同的荧光团修饰的靶向mNP。通过对摄取率的量化，分数 可以测量结合剂的浓度。有大量的体内荧光断层成像设备资源，请访问 达特茅斯，允许使用MicroCT、超声波、高场MRI和全身MRI扫描仪进行测量， 在过去的十年中，软件、硬件和动物研究方面的大量专业知识已经发展到 允许常规应用此方法。该项目将调整现有的硬件/软件以与 并对该程序开发的mNP进行了研究。 MSB能够通过评估mNP在体内直接测量mNP的结合能 布朗运动。目前还没有方法能够做到这一点。MNP在每个位置的平均结合能 位置将被测量。来自OFS的平均结合能和束缚能将提供 估计结合到细胞表面和囊泡中的mNP的比例。体内和体外研究将是 用于验证结合的特异性，以及相对于MSB信号的定位。 这一技术平台是一种全新的、独特的方法来量化体内的mNP结合，并且 这里开发的工具可用于广泛的mnp目标评估。在内部开发时 程序，我们可以将该技术用于其他MNP目标构造，以帮助评估 体内靶向的相对成功，并跟踪随时间的结合状态。它的终极价值在于 技术将允许在临床治疗中纵向跟踪体内结合，从而提供 用于患者特定治疗的定制绑定剂量计。

PROJECT SUMMARY (See instructions); The aim of this project is to develop new methodology to quantitatively image targeted magnetic nanoparticle (mNP) distribution in vivo in each microscopic compartment to provide mechanistic insights into where targeted mNPs collect and how targeted delivery might be improved. The project will combine two novel synergistic methods, optical ratiometric fluorescence spectroscopy (OFS) and magnetic spectroscopy of mNP Brownian motion (MSB), to quantify bulk concentration uptake and then quantify the level of specific binding to the target ligands in vivo, focusing initially on HER2/neu targeted mNPs which can be internalized when bound. Both measurement techniques proposed here have been uniquely developed at Dartmouth and we have considerable existing resources and expertise that can be leveraged. The global distribution and the bound fraction will be measured with OFS by injection of non-targeted and targeted mNP decorated with different fluorophores. Through quantification of the ratio of uptake, the fraction of bound agent can be measured. There is a large resource of in vivo fluorescence tomography apparatus at Dartmouth, allowing measurement with MicroCT, ultrasound, high field MRI and whole body MRI scanners, and substantial expertise in software, hardware and animal studies have developed in the past decade to allow routine application of this approach. The project will adapt existing hardware/software to couple with magnetic spectroscopy and study the mNP developed by this program. MSB has the ability to measure the binding energies ofthe mNP directly in vivo by evaluating the mNP Brownian motion. Currently no methods are able to do this. The average binding energy for the mNP at each position will be measured. The average binding energy and the bound faction from OFS will provide estimates ofthe fraction of mNP bound to the cell surface and in vesicles. In vivo and ex vivo studies will be used to validate the specificity of the binding, as well as the localization relative to the MSB signal. This technology platform is a fundamentally new, unique way to quantify mNP binding in vivo, and the tools developed here can be utilized in a wide range of mNP targeting assessments. When developed within the program, we can make the technology available for other mNP targeting constructs to help assess the relative success of targeting in vivo, and track the binding status over time. The ultimate value of this technology would be to allow longitudinal tracking of binding in vivo in clinical treatments, thereby providing a custom binding dosimeter for use in patient-specific treatments.