High Accurate Micro-Orifice Resistance Assay with Programmable Aggregation–Dispersion Switching Enabled by an Entropy-Driven Circuit

Rapid and sensitive detection of pathogenic bacteria remains challenging due to insufficient signal gain and uncontrollable signal transduction strategies that are easily perturbed by environmental fluctuations. Here, we present a micro-orifice resistance biosensor that converts a molecular circuit-programmed aggregation-dispersion transition into a quantifiable electrical signal. Entropy-driven circuit acts as the core regulatory module, maintaining signal probes in a stable low-entropy aggregated state while providing a built-in, target-responsive pathway for controlled dispersion. Upon recognition of bacterial DNA, the molecular circuit shifts toward a higher-entropy dissociated state, releasing dispersed microparticles that generate distinct resistance pulse signatures. To enhance initiator availability and strengthen the molecular circuit activation, a hairpin-free bridge-hybridization chain reaction (nHCR) supplies complementary upstream amplification without relying on enzymatic processes. The sensor achieves a limit of detection of 27 CFU/mL for Listeria monocytogenes in an amplification-free manner, comparable to quantitative PCR in real sample analysis. Our molecular circuit-programmed dispersion switching, supported by dual amplification, offers a robust and scalable strategy for rapid pathogen analysis.