Comparison of commercial exosome isolation kits for circulating exosomal microRNA profiling
Comparison of commercial exosome isolation kits for circulating exosomal microRNA profiling
复制标题
用于循环外泌体 microRNA 分析的商业外泌体分离试剂盒的比较。
DOI：
10.1007/s00216-018-1052-4
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发表时间：
2018-06-01
影响因子：
4.3
通讯作者：
Zhang, Chunni
中科院分区：
文献类型：
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作者：
Ding, Meng;Wang, Cheng;Zhang, Chunni
循环外泌体microRNAs （miRNAs）是有价值的生物标志物候选者；然而，关于循环外泌体miRNA分析的商用试剂盒的特性和相互一致性的信息很少。在这里，我们分别分析了四种常用的外泌体miRNA分析商用试剂盒的优缺点，以及它们在血清和/或血浆样品中的应用。采用纳米观察和Western blotting对分离的外泌体的效率和纯度进行评价。在我们的条件下，分离颗粒的大小分布是合适的（40-150 nm）， ExoQuick™外泌体沉淀溶液（EXQ）产生的外泌体产量相对较高。然而，白蛋白杂质在所有四种试剂盒中普遍存在，血清或血浆的总外泌体分离（TEI）获得了相对纯净的分离。我们进一步将Illumina测序与RT-qPCR相结合，以确定这些试剂盒对miRNA分析的能力。试剂盒之间的外泌体miRNA谱和特异性miRNA有显著相关性，但因方法不同而存在差异。exoRNeasy血清/血浆Midi试剂盒（EXR）和EXQ在特异性外泌体mirna恢复方面表现更好。EXR、TEI、EXQ和RIBO™外泌体分离试剂（REI）特异性miRNA测定的测定内CVs分别为0.88-3.82、1.19-3.77、0-2.70和1.23-9.11%。在每个试剂盒中，血清比血浆产生更高丰度的外泌体和外泌体mirna，但有更多的白蛋白杂质。总之，我们的数据为循环外泌体mirna的疾病生物标志物鉴定方法提供了一些有价值的指导。循环外泌体microRNAs （miRNAs）是有价值的生物标志物候选者；然而，关于循环外泌体miRNA分析的商用试剂盒的特性和相互一致性的信息很少。在这项研究中，我们比较了四种常用的市售外泌体mirna提取试剂盒，并分析了每种试剂盒的优缺点及其在血清和/或血浆样品中的应用
Circulating exosomal microRNAs (miRNAs) are valuable biomarker candidates; however, information on the characterization and mutual agreement of commercial kits for circulating exosomal miRNA profiling is scarce. Here, we analyzed the advantages and weaknesses of four commonly used commercial kits for exosomal miRNA profiling and their application to the sample of serum and/or plasma, respectively. NanoSight and Western blotting were conducted to evaluate the efficiency and purity of the isolated exosomes. In our conditions, the size distribution of the isolated particles was appropriate (40–150 nm), and ExoQuick™ Exosome Precipitation Solution (EXQ) generated a relatively high yield of exosomes. Nevertheless, albumin impurity was ubiquitous for all the four kits, and Total Exosome Isolation for serum or plasma (TEI) yielded a relatively pure isolation. We further performed Illumina sequencing combined with RT-qPCR to determine the ability of these kits for miRNA profiling. There was significant correlation of the exosomal miRNA profile and specific miRNAs between kits, but with differences depending on methods. exoRNeasy Serum/Plasma Midi Kit (EXR) and EXQ performed better in the specific exosomal miRNAs recovery. Intraassay CVs for specific miRNA measurement were 0.88–3.82, 1.19–3.77, 0–2.70, and 1.23–9.11% for EXR, TEI, EXQ, and RIBO™ Exosome Isolation Reagent (REI), respectively. In each kit, serum yielded a higher abundance of exosomes and exosomal miRNAs than plasma, yet with more albumin impurity. In conclusion, our data provide some valuable guidance for the methodology of disease biomarker identification of circulation exosomal miRNAs.Graphical abstractCirculating exosomal microRNAs (miRNAs) are valuable biomarker candidates; however, information on the characterization and mutual agreement of commercial kits for circulating exosomal miRNA profiling is scarce. In this study, we compared four commonly used commercially available kits for exosomal miRNAsextraction and analyzed the advantages and weaknesses of each kit and their application to the sample ofserum and/or plasma