High-Throughput Purification and Quantification of Bionanoparticles on Capillary-Channeled Fiber Stationary Phases

在化学测量和成像项目的支持下，克莱姆森大学的R. Kenneth Marcus教授和他的团队正在准备、表征和实施一种独特的纤维毛细管通道聚合物（C-CP），以促进生物和合成纳米颗粒的分离、纯化和定量。靶系统包括外泌体，一类细胞外囊泡（ev），它们是细胞内通讯的组成部分，因此参与正常的内稳态功能，但也参与异常生物功能的进化；因此，在此进行的研究对生物医学科学有长期的影响。研究中的应用在聚酯C-CP纤维柱上采用疏水相互作用色谱（HIC）协议，该系统旨在探索基础生物化学的各个方面。研究的总体目标是产生高效、选择性和经济的分离。这项工作是跨学科的，涉及化学、生物工程、数学和材料科学研究小组之间的合作努力。如果成功，该项目可能会在南卡罗来纳州的纺织和生物制药行业带来新的商业风险。与发展多元化劳动力相关的努力包括将研究课题整合到相关课程中。绝大多数生物和合成纳米颗粒的分离/纯化策略涉及物理手段，如超离心、超滤和粒径排除方法。使用C-CP纤维来影响这些分离取决于基于化学功能区分纳米颗粒的能力，其相互作用强度部分取决于颗粒大小。C-CP光纤平台可以实现多种格式，包括自旋向下微移管尖端和微孔柱。因此，可以解决基础生物化学，临床分析和过程分析化学中的相关问题。通过多角度光散射（MALS）在线粒度测定提供了洗脱的纳米颗粒/囊泡的定性和定量信息。采用固定化配体的表面修饰可以靶向纳米颗粒的选择性捕获，并辅以选择性免疫标记策略进行物种特异性分析。多维分离为复杂系统中纳米颗粒的有效表征提供了另一个创新方面。在所有情况下，性能指标都将与商业专栏和标准方法中提供的性能指标进行比较。实用方法的发展将通过在微观水平上理解物理化学过程的基础研究得到加强。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

With support from the Chemical Measurement and Imaging Program in the Division of Chemistry, Professor R. Kenneth Marcus and his group at Clemson University are working to prepare, characterize, and implement a unique format of fiber capillary-channeled polymers (C-CP) to facilitate the isolation, purification and quantification of biological and synthetic nanoparticles. Target systems include exosomes, a class of extracellular vesicles (EVs) that are integral components in intracellular communication, and thus are involved in normal homeostatic function, but also in the the evolution of abnormal biological function; thus there are long term implications of the studies being performed herein for biomedical science. The applications under study employ a hydrophobic interaction chromatography (HIC) protocol on polyester C-CP fiber columns, a system designed to probe aspects of fundamental biochemistry. The overall goal of the studies is to produce efficient, selective, and economical separations. This work is interdisciplinary, involving collaborative efforts among research groups in chemistry, bioengineering, mathematical, and materials sciences. If successful, the project may result in new commercial ventures in the textile and biopharmaceutical industries in South Carolina. Efforts related to developing a diverse workforce include integration of research topics into relevant coursework.The vast majority of separation/purification strategies for biological and synthetic nanoparticles involve physical means such as ultracentrifugation, ultrafiltration, and size-exclusion methods. Use of C-CP fibers to affect these separations rests on the ability to differentiate nanoparticles based upon chemical functionality, whose interaction strengths are determined in part based on particle size. The C-CP fiber platform can be implemented across a number of formats including spin-down micropipette tips and microbore columns. As a result, relevant problems in fundamental biochemistry, clinical analysis, and process analytical chemistry can be addressed. In-line particle sizing via multi-angle light scattering (MALS) provides both qualitative and quantitative information about eluted nanoparticles/vesicles. Surface modifications employing immobilized ligands can target selective capture of nanoparticles, complemented by selective immunolabelling strategies for species-specific assays. Multidimensional separations offer another innovative aspect for efficient characterization of nanoparticles from complex systems. In all cases, performance metrics will be compared to those available from commercial columns and standard methods. Development of practical methods will be augmented with fundamental studies directed at understanding physico-chemical processes at the microscopic level.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.