Streptavidin-functionalized terahertz metamaterials for attomolar exosomal microRNA assay in pancreatic cancer based on duplex-specific nuclease-triggered rolling circle amplification
Streptavidin-functionalized terahertz metamaterials for attomolar exosomal microRNA assay in pancreatic cancer based on duplex-specific nuclease-triggered rolling circle amplification
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基于双链体特异性核酸酶触发滚环扩增的链霉亲和素功能化太赫兹超材料用于胰腺癌阿托摩尔外泌体 microRNA 检测
DOI：
10.1016/j.bios.2021.113314
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发表时间：
2021-05-21
影响因子：
12.6
通讯作者：
Fu, Weiling
中科院分区：
文献类型：
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作者：
Zhan, Xinyu;Yang, Sha;Fu, Weiling
外泌体microRNA（miRNA）是一种很有前途的液体活检的非侵入性生物标志物。本文中，我们制造了太赫兹（THz）超材料生物传感器，其包括由环形凹槽包围的金（Au）盘阵列，用于基于THz特异性核酸酶（DSN）触发的滚环扩增（RCA）的外泌体miRNA测定。在这种策略中，靶miRNA被固定在磁珠（MB）上的探针P0捕获;然后在DSN的作用下重复释放引物P1，DSN作为随后利用生物素-dUTP的RCA步骤的高度特异性引发剂。在靶回收和核酸扩增后，生物素化的扩增产物被链霉亲和素（SA）官能化的THz超材料捕获，并进一步缀合至SA修饰的AuNP，其允许形成SA-生物素化的RCA产物AuNP的三聚体复合物。复杂的群体与目标miR-21的起始浓度成比例，导致太赫兹超材料的共振峰的红移。这种生物传感器可以导致高度特异性和灵敏度的检测与一个碱基的错配歧视和检测限（LOD）低至84 aM。胰腺癌患者和健康对照之间临床血浆样本中外泌体miR-21定量的频率变化存在显着差异。太赫兹超材料的频率偏移与逆转录-聚合酶链反应（RT-PCR）结果一致，说明了我们的分析在真实的临床样本中的适用性和准确性。
Exosomal microRNA (miRNA) is a promising non-invasive biomarker for liquid biopsies. Herein, we fabricated a terahertz (THz) metamaterial biosensor that comprises an array of gold (Au) discs surrounded by annular grooves for exosomal miRNA assays based on duplex-specific nuclease (DSN)-triggered rolling circle amplification (RCA). In this strategy, the target miRNA is captured by a probe P0 immobilized on magnetic beads (MBs); it then repeatedly releases a primer P1 under the action of DSN, which acts as a highly specific initiator of the subsequent RCA step utilizing biotin-dUTP. After target recycling and nucleic acid amplification, the biotinylated amplification products were captured by the streptavidin (SA)-functionalized THz metamaterials, and further conjugated to SA-modified AuNPs that permit formation of a trimeric complex of SA-biotinylated RCA productsAuNP. The complex population scales with the starting concentration of the target miR-21, resulting in a red shift of the resonance peak of the THz metamaterials. This biosensor can lead to highly specific and sensitive detection with one-base mismatch discrimination and a limit of detection (LOD) down to 84 aM. Significant distinctions are seen in the frequency shifts for exosomal miR-21 quantitation in clinical plasma samples between pancreatic cancer patients and healthy controls. The frequency shifts of the THz metamaterials are consistent versus the reverse transcription-polymerase chain reaction (RT-PCR) results, illustrating the applicability and accuracy of our assay in real clinical samples.