Combined determination of copper ions and β-amyloid peptide by a single ratiometric electrochemical biosensor

Copper ions (Cu²⁺) play a critical role in biological processes and are directly involved in β-amyloid peptide (Aβ) aggregation, which is responsible for the occurrence and development of Alzheimer's disease (AD). Therefore, combined determination of Cu²⁺ and Aβ in one analytical system is of great significance to understand the exact nature of the AD event. This work presents a novel ratiometric electrochemical biosensor for the dual determination of Cu²⁺ and Aβ_1-42. This unique sensor is based on a 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulphonate) (ABTS) and poly(diallyldimethylammonium chloride) (PDDA)-bi functionalized single-walled carbon nanotubes (ABTS-PDDA/CNTs) composite. The inclusion of ABTS not only enhanced the sensitivity, but it also acted as an inner reference molecule to improve detection accuracy. The specific recognition of Cu²⁺ was realized by neurokinin B (NKB) coatings on the ABTS-PDDA/CNTs surface to form a [Cu^II(NKB)₂] complex with Cu²⁺. The ABTS-PDDA/CNTs-NKB modified electrode also displayed an excellent electrochemical response toward the Aβ_1-42 monomer, when a certain amount of the Aβ_1-42 monomer was added to Cu²⁺-contained PBS buffer, which was due to the release of Cu²⁺ from the [Cu^II(NKB)₂] complex through Aβ binding to Cu²⁺. Meanwhile, our work showed that Cu²⁺ bound Aβ_1-42 was concentration-dependent. Consequently, the presented electrochemical approach was capable of quantifying two important biological species associated with AD by one single biosensor, with the detection limits of 0.04 μM for Cu²⁺ and 0.5 ng mL^-1 for Aβ_1-42, respectively. Finally, the ratiometric electrode was successfully applied for monitoring Cu²⁺ and Aβ_1-42 variations in plasma and hippocampus of normal and AD rats.