Research Projects

Alzheimer’s disease (AD) is a devastating neurodegenerative disorder affecting millions worldwide, with its prevalence increasing due to aging populations and the lack of effective treatments. Early diagnosis is crucial for timely intervention, potentially slowing disease progression and improving patient outcomes. A promising area of research involves identifying reliable biomarkers for early-stage AD, particularly phosphorylated tau (p-tau) proteins and tau oligomers found in exosomes derived from cerebrospinal fluid (CSF) and blood. These tau species, especially those phosphorylated at Thr-181 and Ser-396, are closely associated with AD pathology. However, current diagnostic tools face significant limitations in detecting and characterizing these biomarkers with high specificity and sensitivity. Traditional immunoassays struggle to differentiate between tau monomers, oligomers, and larger aggregates, providing an incomplete view of the disease. This research aims to develop a novel diagnostic approach combining nanopore technology with high-affinity single-chain antibodies (nanobodies) to address these limitations. Nanopore technology offers zeptoliter (10⁻²¹ L) sensitivity, enabling single-molecule characterization of tau proteins and their aggregates. By applying an electric field, nanopores can analyze tau variants in real-time, distinguishing between monomers and oligomers and providing detailed information on their size, shape, and net charge. The proposed approach uses unique combination of nanopore sensing and nanobodies to discriminate different tau aggregates and quantify their abundance with high specificity. This method also allows for the detection of physiologically relevant post-translational modifications (PTMs), such as Thr-181 and Ser-396 phosphorylation, which are key biomarkers of AD. Functionalizing nanopores with antibodies specific to these PTMs will reveal site-specific phosphorylation, providing deeper insights into disease status and progression. Our research plan is structured into three specific tasks to optimize, validate, and apply the nanopore-based bioassay for tau protein detection in biological samples: Objective 1: Optimization of Exosome Isolation and Nanopore-Assay Configuration 1.1: Exosome Isolation Optimization 1.2: Nanopore-Assay Configuration Objective 2: Assay Robustness and Performance Evaluation 2.1: Assay Robustness Validation 2.2: Performance Metrics Evaluation Objective 3: Application of Optimized Bioassay to Biological Samples 3.1: Clinical Sample Analysis 3.2: Diagnostic Performance Evaluation This innovative approach offers a highly specific and sensitive diagnostic tool that can accurately characterize tau species in AD samples. It has the potential to enable early-stage AD diagnosis, therapeutic monitoring, and assessment of emerging treatments.