ac8b05920_si_001.pdf (1.27 MB)

Metastable Dumbbell Probe-Based Hybridization Chain Reaction for Sensitive and Accurate Imaging of Intracellular-Specific MicroRNAs In Situ in Living Cells

Download (1.27 MB)
journal contribution
posted on 11.03.2019, 00:00 by Jun Chen, Hui-Hui Yang, Wen Yin, Yanfei Zhang, Yingjun Ma, Danping Chen, Yuzhi Xu, Si-Yang Liu, Li Zhang, Zong Dai, Xiaoyong Zou
Sensitive and accurate imaging of intracellular-specific microRNAs (miRNAs) in situ in living cells is seriously challenged by the susceptibility of nucleic acid probes and the low dynamics of the hybridization reaction in cellular environments. Herein, we engineer a set of new metastable dumbbell probes (MxDPs) to overcome these key limitations by concurrently boosting transfection, antidigestibility, assembly dynamics, and nanostructural uniformity. The MxDPs can maintain their stability up to 16 h in living cells and produce uniform and dense DNA nanostructures rapidly (<2 h) and specifically from a hybridization chain reaction (HCR). A sharp signal from the cascade accumulation of fluorescence resonance energy transfer (FRET) further minimizes the effect of system fluctuations. The MxDPs-based HCR (MxDPHCR) method showed identical performance in the analysis of miR-27a in cell lysate and buffer condition and obtained a limit of detection down to 3.2 pM (corresponding to 160 amol per 50 μL), which is 44-fold lower than on conventional hairpin probes. The MxDPHCR method clearly distinguished normal cells from tumor cells and provided more accurate quantitative information on the intracellular-specific miRNAs. The strategy would offer a powerful tool for visualizing and localizing desired nucleic acids in living cells.