Light-Regulated Iontronic Sensing for Resolving the Charge–Steric Trade-Off in the Detection of Exosomal m ⁵ C-Modified miR-21

Exosomal microRNA methylation represents a critical epigenetic modification closely implicated in cancer progression. However, its sensitive detection continues to pose substantial challenges, owing to the inherent coupling between charge accumulation and steric restriction during signal amplification. Here, we report a light-regulated iontronic biosensor for sensitive detection of exosomal m 5 C-modified microRNA-21 (m 5 C-miR-21) by regulating the charge–steric trade-off during hydrogel formation. The DNAzyme selectively recognizes and cleaves m 5 C-miR-21, releasing a trigger strand that initiates the assembly of HP/AuNPs/CaCO 3 probes onto anodic aluminum oxide (AAO) nanochannels. Upon ultraviolet irradiation, o-nitrobenzaldehyde generates H +, which induces CaCO 3 decomposition and Ca 2+ release. The released Ca 2+ subsequently cross-links sodium alginate to form a negatively charged hydrogel in situ on the nanochannel surface. The resulting hydrogel increases local charge density and promotes ion enrichment, while excessive gel growth imposes steric resistance to ion transport. By regulating the irradiation process, the charge–steric trade-off between these two effects is optimized to maximize the ionic current response. When the charge–steric trade-off is properly optimized, the biosensor achieved a linear detection range from 0.1 fM to 0.1 nM with a detection limit of 32 aM. The platform further demonstrated reliable performance in cell lysates and plasma samples, enabling discrimination between colorectal cancer samples and controls. This strategy provides an effective route for iontronic analysis of nucleic acid methylation through light-controlled regulation of charge–steric interplay in nanochannels.