Framework Nucleic Acid-Based Spatial-Confinement Amplifier for miRNA Imaging in Living Cells
Framework Nucleic Acid-Based Spatial-Confinement Amplifier for miRNA Imaging in Living Cells
复制标题
用于活细胞中 miRNA 成像的基于核酸的空间限制放大器框架。
DOI：
10.1021/acs.analchem.1c04866
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发表时间：
2022-02-02
影响因子：
7.4
通讯作者：
Chen, Xiaoqing
中科院分区：
文献类型：
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作者：
Feng, Yinghui;Liu, Qi;Chen, Xiaoqing
活细胞中miRNA的实时原位监测通常被呼吁信号放大器来解决其低丰度的挑战。然而，放大器的不良动力学和来自复杂细胞内环境的潜在干扰阻碍了其在体内的广泛应用。在此，我们报告了一种基于框架核酸（FNA）的非酶空间限制放大器，用于快速可靠的细胞内miRNA成像。放大器由封装在FNA（20 bp立方体）内腔中的局部催化发夹组装（L-CHA）反应器组成。通过将L-CHA的两个探针（H1和H2）整合到DNA链中并将其利用到立方体的对角，L-CHA反应器当然被限制在内部框架中。我们发现，由于FNA的保护，放大器的稳定性显着提高。更重要的是，FNA的空间限制效应可以赋予受限的L-CHA放大器增强的局部试剂浓度（5000倍），从而加速反应速率并提高动态性能（高达14.34倍）。利用这些优点，所提出的放大器可以实现活细胞中miRNA表达水平的准确和有效的监测，并在医学诊断和生物医学研究中具有巨大的潜力。
Real-time in situ monitoring of miRNAs in living cells is often appealed to signal amplifiers to tackle their low abundance challenges. However, the poor kinetics of amplifiers and potential interferences from the complex intracellular environment hamper its widespread applications in vivo. Herein, we report a framework nucleic acid (FNA)-based nonenzymatic spatial-confinement amplifier for rapid and reliable intracellular miRNA imaging. The amplifier consists of a localized catalytic hairpin assembly (L-CHA) reactor encapsulated in the inner cavity of an FNA (a 20 bp cube). The L-CHA reactor is certainly confined to the internal frame by integrating two probes (H1 and H2) of the L-CHA within a DNA strand and harnessing it to the opposite angles of the cube. We find that the stability of the amplifier is remarkably improved due to the protection of the FNA. More importantly, the spatial-confinement effect of the FNA can endow the confined L-CHA amplifier with enhanced local concentrations of reagents (5000-fold), thereby accelerating the reaction rate and improving the dynamic performance (up to 14.34-fold). With these advantages, the proposed amplifier can enable accurate and effective monitoring of miRNA expression levels in living cells and poses great potential in medical diagnostics and biomedical research.