Highly sensitive miRNA-21 detection with enzyme-free cascade amplification biosensor

In this study, we present an enzyme-free fluorescence biosensor for the highly sensitive detection of miRNA-21, a crucial biomarker in clinical diagnosis. Our innovative approach combines catalytic hairpin assembly (CHA) and entropy-driven amplification into a cascade amplification strategy. MicroRNA initiates the catalytic hairpin assembly reaction, liberating the trigger region needed for the entropy-driven amplification reaction. This triggers a series of strand displacement reactions, resulting in the separation of the fluorescence resonance energy transfer pair and an amplified fluorescence signal from FAM. Our cascade amplification strategy achieves ultra-sensitive microRNA detection, with an impressive limit of detection (LOD) of 1.3 fM, approximately 100-fold lower than CHA alone. Additionally, we successfully applied this biosensor for microRNA quantification in human serum and cell lysates, demonstrating its practicality and potential for early diagnosis.