Ultrasensitive Detection of Neurofilament Light in Plasma Using F(Ab')2-Modified Graphene Field-Effect Biosensor.

Neurofilament light (NfL) is a discriminative blood biomarker for many neurological diseases. Current accurate analysis relating to NfL relies on state-of-the-art technologies such as the single-molecule array (Simoa) and immunoprecipitation-mass spectrometry (IP-MS), which require complicated machinery, skilled operational personnel, and well-equipped laboratories. Herein, we demonstrate a robust on-chip graphene field-effect transistor (GFET) biosensing platform for the ultrasensitive detection of NfL. This work utilizes smaller antibody fragments F(ab')2 to mitigate Debye screening and enhance sensing performance, alongside quantitative characterization of 1-pyrenebutyric acid N-hydroxysuccinimide ester (PBASE) surface density to support controlled antibody immobilization. Compared with whole antibody-based GFETs, this F(ab')2-modified GFET platform is shown to achieve a 114% increase in sensitivity, a fivefold improvement in the limit-of-detection (LoD) down to 0.18 pg/mL, and a wide dynamic detection range from 0.18 to 1500 pg/mL, together with good selectivity, stability, and reproducibility. This biosensing platform is validated against Simoa technology for the detection of NfL in clinical plasma samples, yielding a high correlation coefficient of 0.99. These results demonstrate the potential of GFETs for point-of-care diagnosis and the monitoring of neurological diseases in frontline clinical settings, outperforming conventional immunoassays and approaching Simoa sensitivity.