Analysis of Single Particles of Amyloid Beta and α-Synuclein With Seeded Amplification for the Diagnosis of Alzheimer's and Parkinson's Disease.

Research overview

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Neurodegenerative diseases, including Alzheimer's disease (AD) and Parkinson's disease (PD), are characterized by the pathological aggregation of specific proteins such as amyloid beta (Aβ) and α-synuclein, respectively. Early detection of these protein aggregates in biological fluids could facilitate timely diagnosis and therapeutic intervention. This study explores an aggregate amplification approach using the surface-based fluorescence intensity distribution analysis (sFIDA) method to enhance the detection sensitivity of Aβ and α-synuclein seeds. Two amplification strategies were investigated: surface-bound and solution-phase amplification. In the case of Aβ, surface-bound amplification using immobilized Aβ-specific antibodies was tested with synthetic Aβ1-42 seeds and monomeric Aβ1-42 as the substrate. Despite observable aggregation, self-aggregation of the monomeric substrate interfered with seed-dependent amplification, rendering the approach ineffective at physiologically relevant concentrations. Attempts to suppress self-aggregation using blocking peptides, bovine serum albumin, and truncated Aβ11-42 substrates were unsuccessful. Solution-phase amplification followed by surface detection also failed to reliably differentiate seeded aggregation from self-aggregation, indicating that Aβ amplification is unsuitable for diagnostic applications. In contrast, α-synuclein exhibited significantly lower self-aggregation, allowing for more effective seeded amplification. Surface-bound α-synuclein amplification successfully detected synthetic seeds at nanomolar concentrations, while solution-phase amplification further improved sensitivity, enabling detection down to the picomolar range. The method was successfully applied to brain homogenates from transgenic PD model mice, demonstrating the potential for detecting α-synuclein seeds in biological samples. These findings highlight the limitations of Aβ amplification for diagnostic purposes while supporting the feasibility of α-synuclein amplification for PD detection. Future work will focus on optimizing this approach for clinical applications.

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