Development of New Blood Immunoassay and Biomarkers for Tau in the Diagnosis of Alzheimer’s Disease

Post-translational modifications, such as truncation facilitated by proteases and phosphorylation mediated by protein kinases, play pivotal roles in Tau protein function and cellular processes. The detection of distinct Tau forms in plasma has garnered significant interest in the scientific and translational communities. We discovered monoclonal antibodies (mAbs) using the hybridoma technique by immunizing mice with different Tau proteins. Enzyme-linked immunosorbent assay (ELISA), surface plasmon resonance, and Western blot experiments were used to evaluate these mAbs. Following that, we developed immunoassays on ELISA and single-molecule array (Simoa) platforms, respectively. The concentration of Tau fragments in human plasma was detected by Simoa assays to develop novel blood ratio biomarkers. Six mAbs were developed in this study, and on this basis, nine ELISA assays and three Simoa assays were established. The detection limits of Simoa-Assay 1-3 were 0.18, 0.12, and 2.70 pg/mL, respectively. The lower limit of quantification is 0.775, 0.647, and 3.630 pg/mL. The concentrations of three different Tau fragments in plasma were detected by Simoa-Assay 1-3, and the ratios of p-Tau217/t-Tau217 and Tau (1-22)-(187-208)/Tau (1-22)-(210-227) in human plasma are reported for the first time. Our three novel Tau Simoa-assays have the potential to be effective and efficient plasma-based screening tools for Alzheimer's disease (AD). Furthermore, the new ratios p-Tau217/t-Tau217 and Tau (1-22)-(187-208)/Tau (1-22)-(210-227) may be potential novel AD plasma biomarkers.