Three-way junction DNA based electrochemical biosensor for microRNAs detection with distinguishable locked nucleic acid recognition and redox cycling signal amplification

Herein, we present one three-way junction (TWJ) DNA based electrochemical biosensor for sensitive microRNAs detection with locked nucleic acid (LNA) as capture probe and enzyme-free redox cycling signal amplification strategy. LNA owns higher binding affinity and thermal stability compared to normal DNA sequence. In the presence of target microRNA (miRNA-21), novel TWJ structure can be specially triggered with the assistant of methylene blue (MB)tagged DNA signal probe, thus producing an enhanced electrochemical current. Furthermore, a chemical reductant, tris (2-carboxyethyl) phosphine (TCEP), is adopted to lead a relayed electron transport between solution TCEP and TWJ-linked MB tag and amplified whole signal intensity. Based on the selectivity from target-induced LNA/DNA TWJ structure and enzyme-free MB cycling amplification strategy, the designed electrochemical biosensor exhibits sensitive miRNA-21 detection, with a linear range from 100 aM to 100 pM and a low detection limit of 77 aM. In addition, it can successfully discriminate the differences between the miRNA family members, presenting a promising response to the direct determination of target miRNA-21 in real biological fluids. Our work provides an alternative LNA-based electrochemical biosensor way in the area of miRNA diagnostics and clinical analysis without complicated enzyme/nanomaterials-assisted signal amplification.

Automated summary

An electrochemical biosensor based on TWJ DNA with LNA as capture probe and enzyme-free signal amplification strategy was developed for sensitive detection of miRNA-21. The sensor showed a linear range from 100 aM to 100 pM and a low detection limit of 77 aM for miRNA-21 detection, demonstrating promising application in direct determination of target miRNA-21 in biological fluids.