Extracellular vesicles (EVs) are cell‐derived particles with a phospholipid membrane present in all body fluids. Because EV properties change in health and disease, EVs have excellent potential to become biomarkers for diagnosis, prognosis, or monitoring of disease. The only technique capable of detecting, sizing, and phenotyping a million of EVs within minutes is (clinical) flow cytometry. A flow cytometer measures light scattering and fluorescence signals of single EVs. Although these signals contain valuable information about the presence and composition of EVs, the signals are expressed in arbitrary units, which make the comparison of measurement results impossible between instruments and laboratories. Additionally, unintended and undocumented variations in the source, preparation, and analysis of the sample lead to orders of magnitude variations in the measured EV concentrations. Here, we will explain the basics, challenges, and common misconceptions of EV flow cytometry. In addition, we provide an overview of recent standardization initiatives, which are a prerequisite for comparison of clinical data and thus for clinical biomarker exploration of EVs.

中文翻译：

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通过临床流式细胞术可靠地测量细胞外囊泡

细胞外囊泡 (EVs) 是细胞衍生的颗粒，在所有体液中都存在磷脂膜。由于 EV 特性在健康和疾病中会发生变化，因此 EV 具有成为诊断、预后或疾病监测的生物标志物的巨大潜力。唯一能够在几分钟内检测、确定大小和分型 100 万个 EV 的技术是（临床）流式细胞术。流式细胞仪测量单个 EV 的光散射和荧光信号。尽管这些信号包含有关 EV 的存在和组成的有价值的信息，但这些信号以任意单位表示，这使得仪器和实验室之间无法比较测量结果。此外，来源、制备、和样本分析导致测得的 EV 浓度发生数量级变化。在这里，我们将解释 EV 流式细胞术的基础知识、挑战和常见的误解。此外，我们概述了最近的标准化举措，这是比较临床数据以及探索 EV 的临床生物标志物的先决条件。