Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Mahidol University, Bangkok, Thailand.
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Srinakharinwirot University, Nakornnayok, Thailand.
Electrophoresis. 2020 Mar;41(5-6):311-318. doi: 10.1002/elps.201900323. Epub 2020 Jan 20.
Exosome quantification is important for estimation of informative messengers (e.g., proteins, lipids, RNA, etc.) involving physiological and pathological effects. This work aimed to develop a simple and rapid distance-based paper portable device using exosome-capture vesicles (polydiacetylene conjugated with antiCD81) for exosome quantification in cell cultures. This novel concept relied on distinct aggregation of exosomes and exosome-capture vesicles leading to different solvent migration. Distances of the migration were used as signal readouts, which could be detected by naked eye. PDA-antiCD81 as exosome-capture vesicles were optimized (e.g., size, reaction ratio, and concentration) and the paper designs were investigated (e.g., diameter of sample reservoir and lamination layer) to enhance the solvent stop-flow effects. Finally, exosome screening on three cell culture samples (COLO1, MDA-MB-231, and HuR-KO1 subclone) was demonstrated. The method could linearly measure exosome concentrations in correlation with solvent migration distances in the range of 10 -10 particles/mL (R > 0.98) from the cell culture samples. The exosome concentration measurements by the developed device were independently assessed by nanoparticle tracking analysis. Results demonstrated no statistically significant difference (p > 0.05) by t-test. This low-cost and rapid device allows a portable platform for exosome quantification without the requirement of expensive equipment and expertise of operation. The developed device could potentially be useful for quantification of other biomarker-related extracellular vesicles.
外泌体定量对于估计涉及生理和病理效应的有意义信使（例如蛋白质、脂质、RNA 等）非常重要。本工作旨在开发一种简单快速的基于距离的纸便携式设备，该设备使用外泌体捕获囊泡（与抗 CD81 偶联的聚二乙酰亚胺）用于细胞培养中外泌体的定量。这一新概念依赖于外泌体和外泌体捕获囊泡的明显聚集，导致不同的溶剂迁移。迁移距离用作信号读出，可以通过肉眼检测到。优化了 PDA-antiCD81 作为外泌体捕获囊泡的大小、反应比和浓度，并研究了纸设计（例如样品储液器和层压层的直径）以增强溶剂停止流动效应。最后，对三种细胞培养样品（COLO1、MDA-MB-231 和 HuR-KO1 亚克隆）进行了外泌体筛选。该方法可以线性测量细胞培养样品中 10-10 个粒子/mL 范围内与溶剂迁移距离相关的外泌体浓度（R>0.98）。通过纳米颗粒跟踪分析独立评估了开发设备的外泌体浓度测量结果。t 检验结果表明无统计学差异（p>0.05）。这种低成本、快速的设备为外泌体定量提供了一个便携式平台，无需昂贵的设备和操作专业知识。开发的设备可能对其他与生物标志物相关的细胞外囊泡的定量有用。
