Controlled activation of a DNA walker driven by mismatched catalytic hairpin assembly for microRNA imaging in cancer cells and clinical samples

DNA-based molecular machines were recognized as promising tools for microRNA imaging. However, their selectivity and sensitivity were often compromised by premature activation and uncontrolled signal leakage. To address these challenges, an APE1-activated mismatched catalytic hairpin assembly-driven three-dimensional DNA walker (AMDW) was developed. The AMDW system effectively avoided premature activation through a controllable APE1-activation sensing strategy. The mismatched catalytic hairpin assembly (MCHA) served as the driving force, which offered both inherent signal amplification and reduced background signals. Additionally, the incorporation of DNA nanostructure as the three-dimensional track enhanced local concentration, thereby improving walking efficiency. With these advantages, AMDW reached a low detection limit of 0.2 pM for microRNA-21. Moreover, the AMDW successfully differentiated cancer cells and clinical tissue with varying expressions of APE1 and microRNA-21. Therefore, a robust platform for the sensitive and specific detection of intracellular microRNA-21 was established, with promising implications for biomedical research and clinical applications.