AuNPs- Aβ-Ni-HRP sandwich assay: A new sensitive colorimetric method for the detection of Aβ _1-40

Amyloid β-peptide (Aβ) is a key predictor for preclinical diagnosis of Alzheimer's disease (AD) and vascular diseases. In this work, we propose a gold nanoparticle (AuNPs)-Aβ-nickel (Ni)-horseradish peroxidase (HRP) based colorimetric assay for the detection of Aβ_1-40. The consecutive binding of Aβ_1-40 to AuNPs and metal ions is designed and examined for Aβ-specific aggregation of AuNPs and the generation of quantitative colorimetric signals. The affinity of Aβ_1-40 towards various metal ions was studied first, and two metal ions, Cu and Ni, were specifically tested with Metal Ion-Binding Site Prediction (MIB) and High-resolution Electrospray Ionization Mass Spectrometry (HR-ESI-MS). Subsequently, the binding of Aβ_1-40 and AuNPs was examined, and the binding between Aβ-AuNPs and Ni-HRP was finally analyzed by UV-Vis and nano-zetasizer. Based on the characterized dual binding of Aβ_1-40, a colorimetric sandwich assay was developed and the analytical performance of the developed assay has been evaluated with standard solutions and human serum samples. Good linearity within a range from 0 nM to 10 nM was found. The detection limits of 0.22 nM in the standard sample and 0.23 nM in the human serum sample have been demonstrated. The newly developed colorimetric sandwich assay is a short, simple, antibody-free assay and achieves high sensitivity with only 100 μL Aβ_1-40 samples. The assay has immense potential for the detection of Aβ_1-40 in biological or biomedical diagnosis.