Dual-potential ratiometric electrochemiluminescence microRNA bioassay based on inner reference probe and catalytic hairpin assembly

Dual-signal ratiometric biosensors have drawn great attention due to their good reproducibility, reliability, and anti-interference ability in sensitive biosensing. Herein, we propose a dual-potential ratiometric electrochemiluminescence (ECL) biosensor based on an internal reference probe and catalytic hairpin assembly (CHA) signal amplification strategy. The cathodic ECL signal of Au-deposited graphitic carbon nitride nanosheets (Au-g-C 3 N 4 ) was selected as the inner reference, while the anodic ECL response of luminol-H 2 O 2 in solution was used as the reporter signal. After target recognition, Pt nanocluster-labeled DNA strands were introduced onto the surface of the electrode by DNA hybridization, effectively accelerating the production of reactive oxygen species (O 2 •– ) and greatly enhancing the reporter signal with high catalytic activity. The ratio of ECL intensity between the enhanced anodic signal and stable cathodic signal varies with a good linear relationship to the logarithm of microRNA concentration during the detection procedure. Assisted by target-induced CHA signal amplification, this ratiometric ECL biosensor displayed favorable performance for the sensitive detection of microRNA-21 with build-in correction against circumventing fluctuations. The expression level of microRNA-21 in cancer cells is evaluated with favorable reliability and reproducibility and displays great potential and priority in complex biosensing and life assays. • A dual-potential ratiometric ECL biosensor with an internal reference was developed. • Nanoclusters catalysis and CHA process enhance the signal responses. • The internal reference signal corrects the environmental fluctuation. • Sensitive and reliable detection of miRNA-21 can be achieved. • The biosensor shows potential application for miRNA detection in complex samples.