Rationally Anchored Geometry-Controlled DNA Tetrahedral Nanostructures for Attomolar Impedimetric IL-6 Detection

Interleukin-6 (IL-6) is a pleiotropic cytokine whose aberrant elevation drives life-threatening conditions, including sepsis, cytokine storm, and autoimmune disorders, yet existing clinical detection methods demand centralized laboratory infrastructure and multi-hour assay times incompatible with rapid point-of-care decision-making. Here, we report an impedimetric aptasensor built on a programmable tetrahedral DNA nanostructure (TDN) interface anchored to a disposable gold screen-printed electrode (Au-SPE) for the ultrasensitive, label-free detection of IL-6. By systematically varying the number of thiolated base vertices from zero to three, we establish a clear and previously unreported structure-function relationship between multipodal anchoring geometry and charge-transfer resistance modulation: tripodal thiolation yields the most rigid, upright, and electrochemically responsive interface, producing the steepest analytical signal gain upon IL-6 binding at the apex-localised aptamer. Under optimised conditions (pH 7.0, 0.05 µM TDN, MCH passivation), the aptasensor exhibits a linear dynamic range of 0.0001-0.001 pg/mL, a limit of detection of 55 ag/mL , and a sensitivity of 1.55×10 ⁷ Ω (pg mL ^-1 ) ^-1 . Selectivity evaluation against seven physiologically relevant interferents such as TNF-α, BSA, glucose, urea, ascorbic acid, glycine, and cysteine confirms negligible cross-reactivity, with relative responses ranging from 0.57% to 14.35% of the IL-6 signal. Spike-recovery experiments in human serum yield recoveries of 74.0-87.6% (%RSD < 4.5%), and the sensor retains functional activity for at least 21 days under refrigerated storage. This work demonstrates that thiolated vertex number is a critical and tunable design parameter for TDN-based biosensors, offering a modular, disposable platform for sub-femtogram cytokine detection with direct applicability to early sepsis diagnosis and inflammatory disease monitoring.