A Cascaded DNA Circuit in Bead Arrays for Quantitative Single-Cell MicroRNA Analysis

Abstract

Single-cell microRNA (miRNA) analysis helps people understand the causes of diseases and formulate new disease treatment strategies. However, miRNA from a single cell is usually very rare and requires signal amplification for accurate quantification. Here, to amplify the signal, we constructed the cascaded DNA circuits consisting of catalytic hairpin assembly and hybrid chain reaction into the bead array platform, on which the uniformly distributed beads were adopted for miRNA quantification. After exponential signal amplification, a consistent linear correlation between the percentage of fluorescent beads and the copy number of miRNA was detected. The proposed bead array can achieve ultrahigh sensitivity as low as 60 copies of miR-155 and high specificity for distinguishing single nucleotide differences. This method has been successfully applied to the quantitative detection of miRNA in a single cancer cell. The high sensitivity, programmability, and simple workflow of the bead array chip will give a huge advantage in basic and clinical research.

Microwell array chip-based single-cell analysis

“…7c). 92 Any aptamer or probe sequence was compatible with the programmable DNA circuits, which would significantly lower the costs and boost accuracy as well as throughput.…”

“…The cascaded DNA circuits consist of catalytic hairpin assembly and hybrid chain reaction. Reproduced with permission 92. Copyright 2021, American Chemical Society.…”

Recent advances of catalytic hairpin assembly and its application in bioimaging and biomedicine

“…The proposed protocol obtained ultrahigh sensitivity owing to the quadratic amplification elicited by the integration of HCR and CHA, which was also utilized in a lot of other detection methods, such as colorimetry, 62,63 fluorescence, 64,65 electrochemistry 66 and photoelectrochemistry. 67 In addition, Jin and Xu 68 constructed a bead array platform based on a cascaded DNA circuit of CHA-HCR. The platform adopted the ratio of fluorescent beads for quantification, and was applied in single cancer cells, achieving sensitivity down to 60 copies.…”

Label-Free, High-Throughput, Sensitive, and Logical Analysis Using Biomimetic Array Based on Stable Luminescent Copper Nanoclusters and Entropy-Driven Nanomachine

“…Array sensing technologies have become a promising approach to perform high-throughput and multiplex detection of biomarkers (e.g., microRNA) for diseases diagnosis. − Among them, the photoluminescence (PL) array has attracted wide attention on account of its high specificity and rapid response. The traditional PL array usually utilized microplate , and biochip , as substrate, which possessed the disadvantages such as high sample loading volume, limited sensitivity, or complex fabrication process.…”