Characterization of Biological Nanoparticle Subsets

我们开发了用于荧光检测和校准、计数和高通量分选的纳米facs方案，利用纳米facs对血浆和其他生物流体中的EV亚群进行分选。使用多个不同的囊泡群体，我们发现nanoFACS分选能够独特地产生高保真的EV亚群（纯度为95-97%），并具有功能性生物货物（RNA和蛋白质）。我们目前的重点是将这种方法应用于临床重要研究，研究癌症疾病负担、转移潜力和对治疗的反应。作为ACI项目的一部分，我们开发的NanoFACS方法是目前唯一能够基于单个电动汽车特性（而不是批量属性）分析和准备分类功能性电动汽车的方法。此外，我们设计了一类新的标签（分子纳米标签），可以与纳米facs一起使用，不仅可以提高EV表位的检测，而且可以通过流式细胞术进行单分子计数。我们为这个纳米facs项目组装并不断改进的专业仪器、专业知识和生物标本管道在世界上任何其他地方都不存在，这是NCI的一项独特资产。

We developed nanoFACS protocols for fluorescence detection and calibration, for counting, and for high-throughput sorting, to sort EV subsets from plasma and other biofluids by nanoFACS. Using multiple distinct vesicle populations, we find that nanoFACS sorting is uniquely able to produce high fidelity EV subsets (95-97% pure), with functional biological cargo (RNA and protein). Our current focus is exclusively on application of this method for clinically important studies, for the study of cancer disease burden, metastatic potential, and responses to treatment. Our NanoFACS approach that we developed as part of this ACI program is the only method currently able to analyze and preparatively sort functional EVs based on single EV characteristics, rather than bulk attributes. Furthermore, we have devised a new class of labels (Molecular NanoTags) that can be used with nanoFACS, not only to improve the detection of EV epitopes, but also to enable single molecule counting by flow cytometry. The specialized instrumentation, expertise, and biospecimen pipeline that we have assembled and are continuing to improve for this nanoFACS program does not exist anywhere else in the world, is a distinctive asset for NCI.