Single‐Molecule Counting Coupled to Rapid Amplification Enables Detection of α‐Synuclein Aggregates in Cerebrospinal Fluid of Parkinson's Disease Patients

Abstract α‐Synuclein aggregation is a hallmark of Parkinson's disease and a promising biomarker for early detection and assessment of disease progression. The prospect of a molecular test for Parkinson's disease is materializing with the recent developments of detection methods based on amplification of synuclein seeds (e.g. RT‐QuIC or PMCA). Here we adapted single‐molecule counting methods for the detection of α‐synuclein aggregates in cerebrospinal fluid (CSF), using a simple 3D printed microscope. Single‐molecule methods enable to probe the early events in the amplification process used in RT‐QuIC and a precise counting of ThT‐positive aggregates. Importantly, the use of single‐molecule counting also allows a refined characterization of the samples and fingerprinting of the protein aggregates present in CSF of patients. The fingerprinting of size and reactivity of individual aggregate shows a unique signature for each PD patients compared to controls and may provide new insights on synucleinopathies in the future.