An electrochemical biosensor for highly sensitive determination of microRNA based on enzymatic and molecular beacon mediated strand displacement amplification

• A novel biosensor has been developed for highly sensitive and specific detection of target miRNA.
• Strand displacement amplification and enzymatic amplification were used for signal amplification.
• The electrochemical biosensor has been successfully applied to detect miRNA in the serum samples.

In this study, a simple electrochemical biosensor has been established for highly sensitive and specific detection of target microRNA using molecular beacon mediated strand displacement amplification and enzymatic amplification. In the presence of target microRNA, the specific hybridization of microRNA with corresponding domain opens the hairpin structure of a molecular beacon (MB), which triggers strand displacement amplification (SDA) to release nicking DNA triggers. The part of DNA triggers binds to the capture probes immobilized on the gold electrode. Another part of DNA triggers hybridizes with the biotinylated detection probes. The electrochemical signal is obtained by using streptavidin linked to alkaline phosphatase (ST-AP) toward the synthetic enzyme substrate α-naphthyl phosphate (α-NP). Under the optimal conditions, the established biosensor shows high sensitivity and selectivity in a dynamic response range from 50 pM to 10 nM with a detection limit as low as 40 pM (S/N = 3), and good specificity and acceptable reproducibility are achieved for the miRNA-222 detection. This simple and highly efficient biosensor can provide a sensing platform for clinical molecular diagnostics and biomedical research.