Homogeneous electrochemical biosensor for microRNA based on enzyme-driven cascaded signal amplification strategy

Infectious diseases are a long-standing and severe global public health problem. The rapid diagnosis of infectious diseases is an urgent need to solve this problem. MicroRNA (miRNA) plays an important role in the intervention of some infectious diseases and is expected to become a potential biomarker for the diagnosis and prognosis of infectious diseases. It is of great significance to develop rapid and sensitive methods for detecting miRNA for effective control of infectious diseases. In this study, a simple and highly sensitive homogeneous electrochemical method for microRNAs using enzyme-driven cascaded signal amplification has been developed. In the presence of target miRNA, the reaction system produced plenty of MB-labeled single-nucleotide fragments (MB-MF) containing a few negative charges, which can diffuse to the negative surface of the ITO electrode easily, so an obvious electrochemical signal enhancement was obtained. Without the target, MB-HP contains a relatively large amount of negative charges due to the phosphates on the DNA chain, which cannot be digested by the enzyme and cannot diffuse freely to the negatively charged ITO electrode, so only a small signal was detected. The enhanced electrochemical response has a linear relationship with the logarithm of miRNA concentration in the range of 10 fM to 10 nM and the limit of detection as low as 3.0 fM. Furthermore, the proposed strategy showed the capability of discriminating single-base mismatch and performed eligibly in the analysis of miRNA in cell lysates, exhibiting great potential for disease diagnosis and biomedical research. Graphical abstract.