Stimuli-responsive logical gate biosensor based on integrated DNAzyme probe for the discrimination of cancer cells

Combining multiple biomarkers can considerably improve diagnostic accuracy compared to testing individually. Herein, we fabricated an AND logic gate biosensor responsive to stimuli based on a highly integrated DNAzyme sensing probe and the MnO₂ nanosheets nanocarrier to improve the sensitivity and specificity of cancer cell detection. Human apurinic/apyrimidinic endonuclease 1 (APE1) and microRNA (miRNA) act as two "input" biomarkers to execute AND logic gate operation. Logical target recognition, signal transduction and amplification module are integrated into one allosteric DNAzyme probe by target-induced transformation from its locked conformation to active structure, which greatly facilitate the sensing progress. The MnO₂ nanosheets serve as the stimuli-responsive nanocarrier, which can only release the loaded sensing probes and cofactor Mn²⁺ for DNAzyme cleavage after being decomposed by glutathione (GSH). Due to selective release and precise logic operation of the integrated DNAzyme probe, only target cancer cells can exhibit relatively strong fluorescence signal by the specific response of APE1 and miR-222. The proposed logic gate biosensor could not only perform accurate analysis of target cancer cells with high discrimination ratio from normal cells, but also enable visualized detection of miR-222 expression differences in different cancer cell types. This strategy will promote the development of simple and effective biosensors and provide a potential tool for cancer diagnosis and the related biomedical application.