两种分离外泌体方法的比较。

IF 1.3 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY
Journal of Nucleic Acids Pub Date : 2022-10-25 eCollection Date: 2022-01-01 DOI:10.1155/2022/8648373
Mohammad A Aziz, Benedict Seo, Haizal M Hussaini, Merilyn Hibma, Alison M Rich
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引用次数: 1

摘要

外泌体是由细胞释放到细胞外环境的膜结合纳米囊泡。它们携带不同类型的RNA,包括可能对各种疾病诊断有用的mRNA。外泌体的分离一直是一个挑战,因为它们的体积小;因此,本研究比较了两种外泌体分离方法。将Exoquick-TC PLUS™外泌体分离试剂盒(kit)与经典的超离心(UC)外泌体分离方法进行比较。在使用这两种方法获得的样品中,冷冻电子显微镜显示圆形或略微拉长的囊泡,直径从50到150纳米不等,由双层膜分隔。动态光散射导致kit外泌体出现多个峰,而UC外泌体出现单峰。值得注意的是,与kit外泌体相比,UC外泌体中存在更多的总RNA (P < 0.0001)。这反映在随后使用qPCR的mRNA分析中,与kit外泌体相比,UC外泌体的Ct值较低。总之,外泌体表征显示UC和试剂盒样品中都存在外泌体。试剂盒样品呈现额外的峰从DLS可能是由于杂质。总的来说,由于总RNA和mRNA含量较高,UC是后续mRNA分析的更好选择;尽管如此,如果没有超离心机,该试剂盒仍然可以使用,因为所选的五个基因中有四个是使用该试剂盒检测和定量的。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Comparing Two Methods for the Isolation of Exosomes.

Comparing Two Methods for the Isolation of Exosomes.

Comparing Two Methods for the Isolation of Exosomes.

Comparing Two Methods for the Isolation of Exosomes.

Exosomes are membrane-bound nanovesicles released by cells into their extracellular environment. They carry different types of RNA including mRNA which may be useful in the diagnosis of various diseases. Exosome isolation has been a challenge because of their small size; therefore, two exosome isolation methods were compared in this study. The Exoquick-TC PLUS™ exosome isolation kit (kit) was compared with the classic ultracentrifugation (UC) method for exosome isolation. In samples obtained using both methods, cryo-electron microscopy showed round or slightly elongated vesicles with diameters ranging from 50 to 150 nm and delimited by a bilayered membrane. Dynamic light scattering resulted in multiple peaks for kit exosomes, whereas a single peak was observed for UC exosomes. Significantly, more total RNA was present in UC exosomes in contrast to kit exosomes (P < 0.0001). This was reflected in subsequent mRNA analysis using qPCR, where UC exosomes had lower Ct values compared to kit exosomes. In conclusion, exosome characterization revealed the presence of exosomes in both UC and the kit samples. The kit samples presented additional peaks from DLS which might be due to impurities. Overall, due to a higher total RNA and mRNA content, UC is a better option for subsequent mRNA analysis; nevertheless, the kit can still be used if an ultracentrifuge is not available as four out of the five genes selected were detected and quantified using the kit.

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来源期刊
Journal of Nucleic Acids
Journal of Nucleic Acids BIOCHEMISTRY & MOLECULAR BIOLOGY-
CiteScore
3.10
自引率
21.70%
发文量
5
审稿时长
12 weeks
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