MicroRNA-21 electrochemiluminescence biosensor based on Co-MOF–N-(4-aminobutyl)-N-ethylisoluminol/Ti₃C₂T_x composite and duplex-specific nuclease-assisted signal amplification

A novel electrochemiluminescence (ECL) biosensor for the determination of microRNA-21 (miRNA-21) was developed, based on a hybrid luminescent Co-MOF-ABEI/Ti₃C₂T_x composite as an ECL luminophore combined with a duplex-specific nuclease (DSN)-assisted signal amplification strategy. The synthesized Co-MOF-ABEI/Ti₃C₂T_x composite carrying N-(4-aminobutyl)-N-ethylisoluminol (ABEI) exhibited strong and stable ECL in the presence of reactive oxygen species (ROS). The ECL biosensor was fabricated by adsorbing Co-MOF-ABEI/Ti₃C₂T_x onto a glassy carbon electrode and covalently coupling the probe DNA onto the surface of the Co-MOF-ABEI/Ti₃C₂T_x-modified electrode. In the presence of the target miRNA-21, the DSN selectively cleaved the complementary DNA section (S1) to miRNA-21, resulting in the release of the transduction section (S2) and the reuse of miRNA-21 in the subsequent amplification cycle. The interaction of the stem-loop structure of the probe DNA with the Co-MOF-ABEI/Ti₃C₂T_x-modified glassy carbon electrode with S2 strands led to the opening of the annular part of the probe DNA. Then, the opened guanine (G)-rich sequences of probe DNA were exposed and folded into a hemin/G-quadruplex DNAzyme in the presence of hemin. The catalysis of H₂O₂ to ROS by the hemin/G-quadruplex DNAzyme significantly enhanced ECL intensity, and this intensity was logarithmically proportional to the concentration of target miRNA-21 between 0.00001 and 10 nM, having a limit of detection of 3.7 fM. The designed ECL biosensor can detect miRNA-21 extracted from HeLa cells, indicating its promising application in clinical diagnosis and disease prognosis analysis.