Spectroscopic Quantification of Ligand Binding in Vivo

项目总结（见说明）; 本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)在每个微观隔室中的体内分布，以提供对其中 目标mNP收集和如何提高目标交付。该项目将联合收割机 协同方法，光学比率荧光光谱（OFS）和磁光谱， mNP布朗运动（MSB），以量化本体浓度摄取，然后量化特异性抗体的水平。 在体内与靶配体结合，最初集中在可以内化的HER 2/neu靶向mNP上 当绑定。这里提出的两种测量技术都是在达特茅斯独特开发的， 我们拥有大量现有资源和专门知识，可以加以利用。 将通过注射非靶向和非靶向药物，用OFS测量总体分布和结合分数。 用不同荧光团修饰的靶向mNP。通过定量摄取率， 可以测量结合剂的量。存在大量的体内荧光断层摄影设备资源， 达特茅斯，允许测量与微型CT，超声，高场MRI和全身MRI扫描仪， 在过去的十年里，软件、硬件和动物研究方面的大量专业知识得到了发展， 允许这种方法的常规应用。该项目将调整现有的硬件/软件以与 磁谱和研究由该计划开发的mNP。 MSB能够通过对mNP的评价，在体内直接测量mNP的结合能 布朗运动目前没有任何方法能够做到这一点。mNP的平均结合能在每个 位置将被测量。平均结合能和来自OFS的结合分数将提供 估计结合到细胞表面和囊泡中的mNP的分数。将进行体内和离体研究。 用于验证结合的特异性以及相对于MSB信号的定位。 该技术平台是一种全新的、独特的体内定量mNP结合的方法， 这里开发的工具可用于广泛的移动自然人目标评估。当开发在 该计划，我们可以使其他mNP靶向结构的技术，以帮助评估 体内靶向的相对成功，并随时间跟踪结合状态。这件事的最终价值 该技术将允许在临床治疗中纵向追踪体内结合，从而提供一种 用于患者特定治疗的定制结合剂量计。

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.