School of Ophthalmology & Optometry, School of Biomedical Engineering, Wenzhou Medical University, Wenzhou, Zhejiang 325035, China.
Department of Preventive Medicine, Wenzhou Medical University, Wenzhou, Zhejiang 325035, China.
Anal Chem. 2020 Jun 2;92(11):7493-7499. doi: 10.1021/acs.analchem.9b05675. Epub 2020 Apr 13.
Isolation and purification of extracellular vesicles (EVs) from plasma is essential to understand the EV circulation mechanism and discover biomarkers for the early detection of diseases. However, the size range of lipoprotein particles such as high density lipoprotein (HDL), low density lipoprotein (LDL), and very low density lipoprotein (VLDL) overlap that of EVs, making it difficult to remove lipoproteins from EVs. Here, we propose a method for the high efficiency separation of EVs in plasma using agarose gel electrophoresis based on their differences in size and zeta potential properties. Electrophoresis track assays revealed that EVs propagate more slowly than HDL but more quickly than LDL and VLDL in 1% agarose gel with pH 7.4 Tris-Acetate-EDTA (TAE) buffer. The size and morphology of the electrophoresis-recovered products were characterized to be consistent with typical EVs. In addition, the biological function of recovered EVs was investigated with cell uptake tests. The feasibility of this method was further verified with human plasma samples. In summary, this technique has the potential to become a convenient and efficient approach for high-purity EV separation.
从血浆中分离和纯化细胞外囊泡 (EVs) 对于了解 EV 循环机制和发现疾病早期检测的生物标志物至关重要。然而，脂蛋白颗粒（如高密度脂蛋白 (HDL)、低密度脂蛋白 (LDL) 和极低密度脂蛋白 (VLDL)）的大小范围与 EVs 重叠，因此很难从 EVs 中去除脂蛋白。在这里，我们提出了一种使用基于琼脂糖凝胶电泳的方法，根据其大小和 ζ 电位特性的差异，从血浆中高效分离 EVs。电泳轨迹分析表明，在 pH 值为 7.4 的 Tris-醋酸盐-EDTA（TAE）缓冲液中，1%琼脂糖凝胶中 EVs 的迁移速度比 HDL 慢，但比 LDL 和 VLDL 快。对电泳回收产物的大小和形态进行了表征，结果与典型的 EVs 一致。此外，还通过细胞摄取试验研究了回收 EVs 的生物学功能。用人血浆样本进一步验证了该方法的可行性。总之，该技术具有成为高纯度 EV 分离的便捷高效方法的潜力。
