An Integrated Sequential‐Activated DNA Circuit for Reliable Imaging of Intracellular miRNA and Remote Control of Cellular Function

Abstract DNA circuits‐powered live‐cell imaging technology shows great promise for accurate cancer diagnosis and cellular function regulation, yet often suffers from signal leakage by the complex biological environment, thus limiting their ability to control cellular functions. Herein, a highly integrated sequential‐activated DNA (ISD) circuit is developed, enabling the reliable intracellular microRNA (miRNA) imaging and the programmable regulation of membrane protein distribution for accurate tumor cell identification and remote control of cellular functions. By integrating and simplifying two traditional hybridization chain reaction (HCR) modules (HCR‐1 and HCR‐2) into a compact two‐probe system, signal interference is effectively minimized. In this design, the HCR‐2 module is sequestered within the HCR‐1 module, preventing premature activation and ensuring the ISD circuit responds exclusively to the simultaneous presence of both target inputs. This sequential activation strategy significantly reduces signal leakage, enabling the high‐fidelity intracellular miRNA imaging and the precise differentiation of tumor cells from normal cells. Furthermore, by utilizing membrane proteins as inputs, the ISD circuit achieved the selective modulation of membrane protein distribution, facilitating the remote regulation of downstream cellular functions with high specificity. The ISD circuit demonstrates potential for high‐contrast imaging of diverse intracellular biomolecules, offering a versatile platform for precise cell differentiation and behavior modulation.