Profiling Protein Aggregate Size Using Single-Molecule Array Technology

Area of Science:

• Biochemistry
• Neuroscience
• Biotechnology

Background:

• Protein aggregation is a hallmark of neurodegenerative diseases.
• Current methods for characterizing aggregate size in biological samples are limited.

Purpose of the Study:

• To demonstrate that fluorescence intensity in single-molecule array (Simoa) microwells provides size-dependent information about protein aggregates.
• To apply this novel method to analyze tau aggregate size in Alzheimer's disease.

Main Methods:

• Utilized defined synthetic tau assemblies to correlate aggregate size with Simoa microwell fluorescence intensity (brightness).
• Applied Simoa brightness profiling to human brain homogenates from Alzheimer's disease patients and controls.
• Validated findings using single-molecule super-resolution microscopy.
• Assessed dynamic changes in protein aggregation in a neuronal cell model.

Main Results:

• Increasing aggregate size correlated with higher Simoa microwell brightness.
• Alzheimer's disease brain homogenates showed a shift towards larger tau aggregates compared to controls.
• Simoa brightness profiling detected time-dependent increases in aggregate size in a cell model.
• The method robustly reported population-level shifts in aggregate size distributions.

Conclusions:

• Simoa microwell brightness serves as a valuable proxy for protein aggregate size.
• This repurposed technology offers high-throughput structural insights into protein aggregation in neurodegenerative diseases.
• Simoa brightness profiling enhances the understanding of aggregation dynamics and disease mechanisms.