Electrochemical Determination of Amyloid Beta Biomarker Ratios in Serum Using a Pyr‐NHS‐Functionalized 3D Graphene Foam Electrode

Recent studies have shown that the Aβ42/Aβ40 concentration ratio has emerged as a more reliable and clinically informative biomarker compared to the individual concentrations of Aβ42 and Aβ40. A decline in the circulating Aβ42/Aβ40 ratio is associated with the indication of early AD signs, and this ratio is increasingly considered a key parameter of minimally invasive AD diagnosis. To represent the Aβ42/Aβ40 dynamic, Aβ42/Aβ40 mixtures were prepared in human serum to test with the biosensor functionalized with Pyr‐NHS. Electrochemical differentiation studies with simulated analytical ratios were conducted with control measurements using a paired electrochemical assay format, and all RSDs were found below 5%. The biosensor demonstrated high sensitivity and excellent linearity for Aβ42 ( R 2 = 0.99, LOD: 0.27 fM) and Aβ40 ( R 2 = 0.97, LOD: 0.65 fM) in serum. The stability was tested over 8 days (RSD < 10% over 4 days, RSD < 25% over 8 days). The repeatability experiments showed that the Aβ42 and Aβ40 biosensors had excellent repeatability and reproducibility (RSDs < 5%). This outstanding sensing performance was ascribed to the stable Pyr‐NHS functionalization via noncovalent π‐π stacking, which highly preserves the intrinsic electroconductivity; the 3D graphene foam, which facilitates enhanced conductivity and an increased electrode surface area; and the optimized binding approach.