DNA Molecular Sieve-Assistant Autocatalytic Circuits for High-Precision Cell Discrimination through an Improved Cellular Spatial Resolution

The high-precision discrimination of cancer cell types is crucial for the fundamental understanding of their progressions and accurate clinical prognosis. Resolving microRNAs (miRNAs) and related biomolecules has emerged as a powerful approach to elucidate cell types. However, the cell discrimination via miRNA profiling needs to address two critical challenges: (i) the shared sequence homology and localization of miRNA to precursor microRNA (pre-miRNA) may lead to false-positive signals. (ii) The existence of the typical miRNA expression across various cell lines may impair accurate cell discrimination. Herein, we present a DNA engineering framework screening (DEFENSE) system integrated into an AND-gated circuit for high-precision cell differentiation. The DEFENSE selectively screens target miRNA-21 via size exclusion from its pre-miRNA, greatly improving the recognition rate of miRNA-21. Simultaneously, the circuit features dual gating: an upstream gate regulated by apurinic/apyrimidinic endonuclease 1 (APE1) sites positioned externally to the DNA framework, and a downstream gate governed by miRNA-21 recognition modules integrated within the DEFENSE framework. This design advances the precision of cell discrimination due to the avoiding of the coexistence of biomarkers across different cell types. Moreover, multiple DEFENSE cages interconnect via catalytic hairpin assembly (CHA)-driven Y-structures, assembling into giant networks for high-resolution intracellular miRNA-21 imaging. The proposed platform has achieved high-precision discrimination among cancer cells of MCF-7, HeLa, and normal cell of MCF-10A at varying expression levels of miRNA-21 and APE1. We anticipate that this method can be widely applicable for accurate cancer diagnoses and disease monitoring among a variety of cell types through more types of biomolecule screening.