Comparison of serum exosome isolation methods for microRNA profiling
Comparison of serum exosome isolation methods for microRNA profiling
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DOI：
10.1016/j.clinbiochem.2013.10.020
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发表时间：
2014-01-01
影响因子：
2.8
通讯作者：
Peters, Maire
中科院分区：
文献类型：
--
作者：
Rekker, Kadri;Saare, Merli;Peters, Maire
目的：外泌体是由大多数细胞分泌的小的膜结合囊泡。外泌体含有各种功能性蛋白质、mRNA和microRNA（miRNA），其可用于诊断和治疗目的。目前，用于血清外泌体分离的标准方法是差异超离心，但正在寻找用于临床环境的替代的、耗时较少且劳动量大的外泌体分离方法。血清外泌体分离方法对获得的miRNA谱的影响尚不清楚。本研究的目的是确定选择的外泌体分离方法在多大程度上影响血清外泌体miRNA profile.Design和方法：通过超离心和ExoQuick沉淀方法从健康个体的血清中分离外泌体。通过真实的时间PCR使用Exigon miRCURY LNA（TM）microRNA Human panel I assays测定375个miRNA的表达谱。结果：虽然在两种分离方法之间观察到外泌体miRNA谱的强相关性，但是在所使用的方法之间鉴定出不同的miRNA水平簇。结论：两种外泌体分离方法均适用于血清外泌体miRNA谱分析。两种方法之间的miRNA模式的差异表明，所观察到的外泌体的miRNA谱受到细胞外囊泡分离方法的轻微影响。(C)2013年加拿大临床化学家协会。爱思唯尔公司出版All rights reserved.
Objectives: Exosomes are small membrane bound vesicles secreted by most cell types. Exosomes contain various functional proteins, mRNAs and microRNAs (miRNAs) that could be used for diagnostic and therapeutic purposes. Currently, a standard method for serum exosome isolation is differential ultracentrifugation, but a search for alternative, less time-consuming and labour extensive exosomal isolation method for use in clinical settings is ongoing. The effect of serum exosome isolation method on obtained miRNA profile is not yet clear. The aim of this study was to determine to which extent selected exosome isolation methods influence the serum exosomal miRNA profile.Design and methods: Exosomes were isolated from blood serum of healthy individuals by ultracentrifugation and ExoQuick Precipitation methods. The expression profile of 375 miRNAs was determined by real time PCR using Exigon miRCURY LNA (TM) microRNA Human panel I assays.Results: Although a strong correlation of exosomal miRNA profiles was observed between the two isolation methods, distinct clusters of miRNA levels between the used methods were identified. The detected levels of two miRNAs, miR-92a and miR-486-5p, were significantly influenced by the exosome isolation method used.Conclusions: Both exosome isolation methods are suitable for serum exosomal miRNA profiling. Differences found in miRNA patterns between the two methods indicate that the observed exosomal miRNA profile is slightly affected by the extracellular vesicle isolation method. (C) 2013 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.