Target-induced proximity ligation triggers recombinase polymerase amplification and transcription-mediated amplification to detect tumor-derived exosomes in nasopharyngeal carcinoma with high sensitivity
Target-induced proximity ligation triggers recombinase polymerase amplification and transcription-mediated amplification to detect tumor-derived exosomes in nasopharyngeal carcinoma with high sensitivity
复制标题
靶标诱导的邻近连接触发重组酶聚合酶扩增和转录介导的扩增，以高灵敏度检测鼻咽癌中肿瘤来源的外泌体。
DOI：
10.1016/j.bios.2017.11.033
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发表时间：
2018-04-15
影响因子：
12.6
通讯作者：
Zhang, Ge
中科院分区：
文献类型：
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作者：
Liu, Wanli;Li, Jianpei;Zhang, Ge
肿瘤来源的外泌体（TEXs）是肿瘤细胞持续释放到血液中的细胞外囊泡，携带原始肿瘤细胞的特定表面标志物。大量证据表明，TEXs是颇具吸引力的癌症诊断标志物。然而，由于早期肿瘤阶段血液中TEXs浓度极低，对其进行检测颇具挑战性。在此，我们建立了一种名为PLA - RPA - TMA的检测方法，能够高灵敏度、高特异性地检测TEXs。基于两种可识别TEX上生物标志物的邻近连接测定（PLA）探针，我们为特定生物标志物生成了独特的替代DNA信号，该信号通过重组酶聚合酶扩增（RPA）与转录介导扩增（TMA）联用的方式同步进行两次扩增，随后采用基于金纳米粒子的比色法对RPA - TMA反应产物进行定量检测。我们利用来自鼻咽癌（NPC）细胞的TEXs为该方法提供了概念验证证据，其检测限可达10²个颗粒/毫升，并报告了血浆中爱泼斯坦 - 巴尔病毒潜伏膜蛋白1（LPM1）阳性（LMP1⁺，准确度：0.956）和表皮生长因子受体（EGFR）阳性（EGFR⁺，准确度：0.906）的TEXs作为鼻咽癌有效早期诊断生物标志物的检测情况。
Tumor-derived exosomes (TEXs) are extracellular vesicles that are continuously released into the blood by tumor cells and carry specific surface markers of the original tumor cells. Substantial evidence has implicated TEXs as attractive diagnostic markers for cancer. However, the detection of TEXs in blood at an early tumor stage is challenging due to their very low concentration. Here, we established a method called PLA-RPA-TMA assay that allows TEXs to be detected with high sensitivity and specificity. Based on two proximity ligation assay (PLA) probes that recognize a biomarker on a TEX, we generated a unique surrogate DNA signal for the specific biomarker, which was synchronously amplified twice by recombinase polymerase amplification (RPA) coupled with transcription-mediated amplification (TMA), and then the products of the RPA-TMA reaction were quantitatively detected using a gold nanoparticle-based colorimetric assay. We established proof-of-concept evidence for this approach using TEXs from nasopharyngeal carcinoma (NPC) cells, with a detection limit of 102 particles/mL, and reported the measurement of plasma Epstein-Barr virus latent membrane protein 1 (LPM1)-positive (LMP1(+), accuracy: 0.956) and epidermal growth factor receptor (EGFR)-positive (EGFR(+), accuracy: 0.906) TEXs as potent early diagnostic biomarkers for NPC.