A label-free electrochemical aptasensor enables ultrasensitive and specific detection of neurofilament light.

Plasma neurofilament light chain (NfL) has been identified as a promising early-stage Alzheimer's disease (AD) biomarker, reflecting neuroaxonal degeneration at preclinical stages. However, its extremely low concentration in blood presents a major analytical challenge. Although single-molecule array (Simoa) assays provide exceptional sensitivity, they remain costly, instrumentation-intensive, and difficult to implement in decentralized settings. Here, we report the identification and engineering of high-affinity DNA aptamers against NfL using capillary electrophoresis-SELEX. Solution-phase fluorescence polarization was used to evaluate the binding capability of the aptamer candidates and the effect of aptamer truncation. Molecular modelling provided structural insights into aptamer-protein interactions, thereby supporting the rational optimization of aptamer design. The optimized truncated aptamers were integrated onto microfabricated gold electrodes to construct electrochemical aptasensors, exhibiting a Kd value of 1.67 ± 0.47 nM. To further enhance analytical performance, the aptamers were implemented in ion-gated organic electrochemical transistors (iOECTs), yielding an ultrasensitive and highly selective platform capable of detecting NfL in 0.1 × human serum with an ultralow limit of detection of 9 aM. This work establishes a scalable, minimally invasive, cost-effective alternative to antibody-based assays and provides a versatile biosensing strategy for early neurodegenerative disease diagnostics.