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基于核酸酶辅助信号放大的2’-O-甲基修饰分子信标用于高灵敏和特异性外泌体肿瘤microRNA分析
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Abstract:
外泌体是新兴的重要癌症生物标志物。有效检测外泌体和外泌体内容物，特别是microRNA (miRNA)，对于癌症的诊断和治疗是迫切且具有挑战性的。基于双链特异性核酸酶(Duplex specific nuclease, DSN)的特异性识别和消化能力，我们设计了一个2’-O-甲基修饰的分子信标(2’-O-methyl-modified molecular beacon, omMB)，并开发了一个高灵敏度和特异性分析外泌体miRNA的信号放大检测平台。该方法以A375细胞分泌的外泌体为模型，以microRNA-21 (miR-21)为模型miRNA分子，可以检测到低至37.9 pM的miRNA和2 μg/mL的裂解外泌体。同时，与许多已开发的DSN辅助信号放大方法相比，新方法具有较高的特异性，可以区分错配miRNA。总之，这项工作为医学分析、临床应用和疾病诊断中的外泌体检测提供了一种有效的分析策略。
Exosomes are emerging important cancer biomarkers. Effective detection of exosome and exosomal contents, especially microRNA (miRNA), is urgent and challenging for cancer diagnosis and treatment. Based on specific recognition and digestion capacity of duplex specific nuclease (DSN), we designed a 2’-O-methyl-modified molecular beacon (omMB) and proposed an amplified detection platform for highly sensitive and specific analysis of exosomal miRNA. Taking A375 cell secreted exosomes as model targets and microRNA-21 (miR-21) as model miRNA molecules, the proposed method can detect miRNA down to 37.9 pM and 2 μg/mL lysed exosomes. Meanwhile, compared with many developed DSN-assisted amplified methods, the new method has high specificity which can distinguish mismatch miRNA. Overall, this work provides an effective analytical strategy for exosome detection in medical analysis, clinic applications and disease diagnosis.
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