Finnish Red Cross Blood Service. Härkälenkki 13, 01730 Vantaa, Finland.
Finnish Red Cross Blood Service. Härkälenkki 13, 01730 Vantaa, Finland.
J Chromatogr A. 2023 Sep 27;1707:464293. doi: 10.1016/j.chroma.2023.464293. Epub 2023 Aug 9.
Purification of extracellular vesicles for research and therapeutic applications requires updated methodology to address the limitations of traditional ultracentrifugation and other size-based separation techniques. Their downfalls include induced extracellular vesicle aggregation, low yields, poor scalability and one-dimensionality of the separation process, as the size or sedimentation speed of extracellular vesicles is often the only selection criterion. Ion exchange chromatography is a promising alternative or supplementary method candidate, as it offers a different approach for extracellular vesicle separation, which is surface charge. For now, mostly anion exchange chromatography has been evaluated for extracellular vesicle purification, as it successfully relies on the strongly negative surface charge of extracellular vesicles. However, as extracellular vesicles are very complex in their structure, also cation exchange chromatography could be applicable, due to individual cationic domains on the extracellular vesicle surface. Here, we compare anion exchange chromatography to different types of cation exchange chromatography for the purification of platelet extracellular vesicle samples also containing plasma-derived impurities. We found that the choice of resin structure used for cation exchange chromatography is critical for binding platelet extracellular vesicles, as a conventional-type cation exchanger was found to only capture and elute less than 20% of extracellular vesicles. With the tentacle-type resin, it was possible to obtain comparable platelet extracellular vesicle yields (over 90%) with cation exchange chromatography compared to anion exchange chromatography, as well as superior purity, especially when it was combined to conventional cation exchange resin.
为了研究和治疗应用的目的，需要对细胞外囊泡进行纯化，这就需要更新的方法来解决传统的超速离心和其他基于大小的分离技术的局限性。它们的缺点包括诱导细胞外囊泡聚集、产量低、分离过程的可扩展性差和一维性，因为细胞外囊泡的大小或沉降速度通常是唯一的选择标准。离子交换色谱是一种很有前途的替代或补充方法，因为它为细胞外囊泡的分离提供了一种不同的方法，即表面电荷。到目前为止，主要是阴离子交换色谱被用于细胞外囊泡的纯化，因为它成功地依赖于细胞外囊泡强烈的负表面电荷。然而，由于细胞外囊泡在结构上非常复杂，阳离子交换色谱也可能适用，因为细胞外囊泡表面存在个别阳离子域。在这里，我们比较了阴离子交换色谱和不同类型的阳离子交换色谱用于纯化也含有血浆衍生杂质的血小板细胞外囊泡样品。我们发现，用于阳离子交换色谱的树脂结构的选择对于结合血小板细胞外囊泡是至关重要的，因为发现传统型阳离子交换剂只能捕获和洗脱不到 20%的细胞外囊泡。使用触手型树脂，可以通过阳离子交换色谱获得与阴离子交换色谱相当的血小板细胞外囊泡产率（超过 90%），并且纯度更高，尤其是当它与传统的阳离子交换树脂结合使用时。
