Highly Effective Detection of Exosomal miRNAs in Plasma Using Liposome-Mediated Transfection CRISPR/Cas13a.
Highly Effective Detection of Exosomal miRNAs in Plasma Using Liposome-Mediated Transfection CRISPR/Cas13a.
复制标题
DOI：
10.1021/acssensors.2c01683
复制
发表时间：
2023-02
期刊：
影响因子：
8.9
通讯作者：
Junli Zhang;Mengting Guan;Chihong Ma;Y. Liu;Min Lv;Zhenzhong Zhang;Hua Gao;Kaixiang Zhang
中科院分区：
文献类型：
--
作者：
Junli Zhang;Mengting Guan;Chihong Ma;Y. Liu;Min Lv;Zhenzhong Zhang;Hua Gao;Kaixiang Zhang
外泌体miRNAs在癌症生物学中起着重要作用，可能成为癌症诊断的潜在生物标志物。然而，由于外泌体中miRNAs的丰度较低，以易于操作的方式识别和检测疾病相关的外泌体miRNAs仍然是一个挑战。在本文中，我们使用脂质体介导的膜融合策略（MFS）将CRISPR/Cas 13 a转染到外泌体中，称为MFS-CRISPR，直接测量血浆中的外泌体miRNA。使用MFS-CRISPR平台检测外泌体miR-21，我们实现了跨越四个数量级（104-108个颗粒/mL）的线性范围，并且该方法能够检测低至1.2 × 103个颗粒/mL的外泌体miR-21。脂质体介导的MFS可以将荧光信号限制在融合的囊泡中，这可以用于外来体异质性分析。此外，通过测量临床样本对MFS-CRISPR测定进行评估，乳腺癌患者和健康供体的miR-21表达差异显著。由于该方法灵敏度高、操作简单，在肿瘤诊断和治疗监测方面具有广阔的应用前景。
Exosomal miRNAs play a critical role in cancer biology and could be potential biomarkers for cancer diagnosis. However, due to the low abundance of miRNAs in the exosomes, recognizing and detecting disease-associated exosomal miRNAs in an easy-to-operate way remain a challenge. Herein, we used a liposome-mediated membrane fusion strategy (MFS) to transfect CRISPR/Cas13a into exosomes, termed MFS-CRISPR, directly measuring exosomal miRNAs in plasma. Using the MFS-CRISPR platform for detection of the exosomal miR-21, we achieve a linear range spanning four orders of magnitude (104-108 particles/mL) and the method is able to detect the exosomal miR-21 in as low as 1.2 × 103 particles/mL. The liposome-mediated MFS could confine fluorescent signals in fused vesicles, which can be used for exosome heterogeneity analysis. Moreover, MFS-CRISPR assay was evaluated by measuring clinical samples, and the difference of miR-21 expression of breast cancer patients and healthy donors was significant. Because of high sensitivity and simplicity, the proposed method could have promising clinical potential for cancer diagnosis and treatment monitoring.