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Qiuyuan Lin, Zhipeng Huang, Xin Ye, Bin Yang, Xueen Fang, Baohong Liu, Hui Chen, Jilie Kong,
Lab in a tube: Isolation, extraction, and isothermal amplification detection of exosomal long noncoding RNA of gastric cancer,
Talanta,
Volume 225,
2021,
122090,
ISSN 0039-9140,
https://doi.org/10.1016/j.talanta.2021.122090.
(https://www.sciencedirect.com/science/article/pii/S0039914021000114)
Abstract: Tumor-derived exosomes that inherit molecular information on parental cells hold great promise for cancer diagnostics. Currently, two main technical challenges, time-consuming and labor-intensive isolation of exosome and nucleic acid extraction with limited recovery that have restricted the detection of ultralow abundance exosomal nucleic acids. Here, we proposed a simple, efficient and “l(fā)ab in a tube” system for the detection of exosomal nucleic acids, which fully integrated exosomes enrichment using immunomagnetic beads (IMB) (10 min), fast exosomes lysis based on NP-40 lysate (5 min) and sensitive loop-mediated isothermal amplification (LAMP) in a tube. This method was demonstrated by detecting two exosomal long noncoding RNA biomarkers of gastric cancer (HOTTIP and lncRNA-GC1) with a dynamic detection ranging from 300 ng/μL to 10 ng/μL, and the detection limit of LAMP was 10 ng/μL. Additionally, this platform exhibited good performance in the analysis of exosomal HOTTIP RNA directly in human serum samples, which has the potential for detection of low-abundance exosomal nucleic acid biomarkers from cancers.

Keywords: Exosomal long noncoding RNA; Immunomagnetic beads isolation; Nucleic acid extraction; Loop-mediated isothermal amplification (LAMP)

摘要：腫瘤來源的外泌體繼承了親代細胞的分子信息，在癌癥診斷方面具有廣闊的前景。目前，兩個主要的技術挑戰(zhàn)，費時費力的外泌體分離和回收率有限的核酸提取限制了超低豐度外泌體核酸的檢測。在這里，我們提出了一種用于檢測外泌體核酸的簡單、高效和“管中實驗室”系統(tǒng)，該系統(tǒng)完全整合了使用免疫磁珠（IMB）（10 分鐘）的外泌體富集，基于 NP-40 裂解物的快速外泌體裂解（ 5 分鐘）和靈敏的環(huán)介導等溫擴增 (LAMP) 在管中。該方法通過檢測胃癌的兩種外泌體長非編碼RNA生物標志物（HOTTIP和lncRNA-GC1）來證明，動態(tài)檢測范圍為300 ng/μL至10 ng/μL，LAMP的檢測限為10 ng/μL。此外，該平臺在直接分析人血清樣品中的外泌體 HOTTIP RNA 方面表現出良好的性能，具有檢測癌癥中低豐度外泌體核酸生物標志物的潛力。
關鍵詞：外泌體長鏈非編碼RNA；免疫磁珠分離；核酸提??；環(huán)介導等溫擴增 (LAMP)