PDF(Pubmed)
Abstract：
Extracellular vesicles (EVs) have emerged as a promising tool for clinical liquid biopsy. However, the identification of EVs derived from blood samples is hindered by the presence of abundant plasma proteins, which impairs the downstream biochemical analysis of EV-associated proteins and nucleic acids. Here, we employed optimized asymmetric flow field-flow fractionation (AF4) combined with density cushion ultracentrifugation (UC) to obtain high-purity and intact EVs with very low lipoprotein contamination from human plasma and serum. Further proteomic analysis revealed more than 1000 EV-associated proteins, a large proportion of which has not been previously reported. Specifically, we found that cell-line-derived EV markers are incompatible with the identification of plasma-EVs and proposed that the proteins MYCT1, TSPAN14, MPIG6B and MYADM, as well as the traditional EV markers CD63 and CD147, are plasma-EV markers. Benefiting from the high-purity of EVs, we conducted comprehensive miRNA profiling of plasma EVs and nanosized particles (NPs), as well as compared plasma- and serum-derived EVs, which provides a valuable resource for the EV research community. Overall, our findings provide a comprehensive assessment of human blood EVs as a basis for clinical biopsy applications.
摘要：
细胞外囊泡（EV）已成为临床液体活检的有希望的工具。然而,来自血液样本的电动汽车的鉴定受到丰富的血浆蛋白的存在的阻碍，这损害了EV相关蛋白和核酸的下游生化分析。这里,我们采用优化的不对称流场-流动分级(AF4)结合密度垫式超速离心(UC),从人血浆和血清中获得了脂蛋白污染极低的高纯度完整EV.进一步的蛋白质组学分析揭示了1000多种EV相关蛋白，其中很大一部分以前没有报道过。具体来说,我们发现细胞系衍生的EV标记与血浆EV的鉴定不相容，并提出MYCT1，TSPAN14，MPIG6B和MYADM蛋白，与传统的EV标志物CD63和CD147一样，也是血浆EV标志物。受益于电动汽车的高纯度，我们对血浆EV和纳米颗粒（NPs）进行了全面的miRNA分析，以及比较血浆和血清来源的电动汽车，这为电动汽车研究界提供了宝贵的资源。总的来说,我们的研究结果为人类血液EV提供了全面评估,作为临床活检应用的基础.
