No need for beads! Virus-derived biological standards for nanoscale flow cytometry

纳米级流式细胞术（NFC）是一种新兴的变革性技术，旨在分析微小生物颗粒上表达的细胞源性表面标志物的组成，最显著的是病毒和细胞外囊泡（EV）。EV类似于所有细胞类型不断释放的微小气泡，并作为协调生长，分化的通信手段，但也警告其他细胞潜在的危险，如损伤，感染和癌症，帕金森氏症和阿尔茨海默氏症等疾病。分析从血液、唾液和尿液等生物液体中分离出的EV的表面标志物可以提供疾病或感染的早期预警信号。因此，EV表面上的标记物有望成为医学和兽医临床诊断的有价值和强大的工具。尽管NFC能够分析EV表面标记，但由于缺乏合适的商业仪器校准标准和实验控制，医院和研究中心对其的广泛部署和采用受到阻碍。我们已经开发出了模拟电动汽车大小和物理特性的荧光病毒衍生颗粒，从而作为NFC的非常可靠和有效的控制。这些病毒可以持续、廉价和大量地生产。本文提出的工作将进一步改进这些病毒，以满足特定的研究和临床要求。预期的结果是生产10种不同的病毒衍生标准品或对照品，可用于各种研究和临床NFC应用，以准确分析EV表面标志物。我们将通过创建一家致力于制造这些病毒衍生颗粒的初创公司来确保这些病毒衍生颗粒的供应。该初创公司将为NFC仪器制造商和试剂分销商提供病毒衍生标准品，并将其提供给学术和行业客户。对加拿大的经济和社会效益包括培训高素质的人员，以及创建一家初创公司，生产NFC应用的试剂，这反过来将导致新一代用于人类，鱼类和牲畜的疾病和感染的非侵入性诊断测试。

Nanoscale flow cytometry (NFC) is an emerging and transformative technology that is designed to analyze the composition of cell-derived surface markers expressed on minute biological particles, most notably viruses and extracellular vesicles (EVs). EVs resemble tiny bubbles constantly released by all cell types and serve as a means of communication to coordinate growth, differentiation, but also to warn other cells of potential dangers such as injury, infection and diseases such as cancer, Parkinson's and Alzheimer's. Analyzing the surface markers of EVs isolated from biological fluids such as blood, saliva and urine may provide early warning signs of disease or infection. As such, markers on the surface of EVs are poised to become valuable and powerful tools for medical and veterinary clinical diagnostics. Despite NFC being capable of analyzing EV surface markers, its broad deployment and adoption by hospitals and research centers has been hindered by the absence of suitable commercial instrument calibration standards and experimental controls. We have developed fluorescent virus-derived particles that mimic the size and physical properties of EVs, and thereby act as extremely reliable and effective controls for NFC. The viruses can be produced consistently, cheaply and in large quantities. The work proposed here will further improve these viruses to meet specific research and clinical requirements. The anticipated outcome is the production of 10 different virus-derived standards or controls that can be used in various research and clinical NFC applications for the accurate analysis of EV surface markers. We will ensure the supply of theses virus-derived particles through the creation of a start-up company dedicated to making them. The start-up will supply NFC instrument manufacturers and reagent distributors with the virus-derived standards, and they will make them available to their academic and industry clients. Economic and societal benefits to Canada include the training of highly qualified personnel, and the creation of a start-up that will manufacture a reagent for NFC applications which will, in turn, lead to a new generation of non-invasive diagnostic tests for disease and infection for use in humans, fish and livestock.**************