Ultrasensitive Electrochemiluminescence Biosensing Platform for Detection of Multiple Types of Biomarkers toward Identical Cancer on a Single Interface

Electrochemiluminescence (ECL) with high sensitivity and excellent controllability provides a promising approach for ultrasensitive detection of multiple biomarkers. However, the detection for multiple types of biomarkers on a single interface remains a considerable challenge owing to the functional differentiation of different types of biomarkers. Herein, we utilized “on–off–on” switching, target-induced cleavage of peptide, and TdT (terminal deoxynucleoside transferase)-mediated extension successfully constructing a novel ECL biosensor for the ultrasensitive detection of microRNA-141 (miRNA-141) and matrix metalloproteinase-2 (MMP-2). Importantly, the dual biomarkers are related with several identical cancers, which endow the biosensor with diagnostic accuracy and efficiency. In this protocol, target 1 (miRNA-141) first hybridized with probe DNA (pDNA) assembled on CdS QDs modified sensing surface. Afterward, miRNA-141 captured trigger DNA (tDNA) to generate a long ssDNA nanotail via TdT-mediated DNA polymerization. Then the forming ssDNA could capture abundant Fc-peptide-ssDNA conjugates through the hybridization reaction, the ECL intensity quenched significantly due to the efficient quenching effect of Fc to CdS QDs, realizing the ultrasensitive detection of miRNA-141 with a detection limit of 33 aM (S/N = 3). After incubated with target 2 (MMP-2) which specifically cleaved the Fc-peptide-ssDNA conjugates causing the releasing of Fc from the sensing surface, the ECL intensity had an obvious enhancement, achieving the ultrasensitive analysis of MMP-2 with a detection limit of 33 fg·mL –1 (S/N = 3). More importantly, this biosensor also realized the monitoring of biomarkers in different cancer cells and human serum, which indicated that the biosensing system could serve as applicable tools in clinical analysis.