RT-QuIC and Related Assays for Detecting and Quantifying Prion-like Pathological Seeds of α-Synuclein

Various disease-associated forms or strains of α-synuclein (αSyn(D)) can spread and accumulate in a prion-like fashion during synucleinopathies such as Parkinson’s disease (PD), Lewy body dementia (DLB), and multiple system atrophy (MSA). This capacity for self-propagation has enabled the development of seed amplification assays (SAAs) that can detect αSyn(D) in clinical samples. Notably, α-synuclein real-time quaking-induced conversion (RT-QuIC) and protein misfolding cyclic amplification (PMCA) assays have evolved as ultrasensitive, specific, and relatively practical methods for detecting αSyn(D) in a variety of biospecimens including brain tissue, CSF, skin, and olfactory mucosa from synucleinopathy patients. However, αSyn SAAs still lack concordance in detecting MSA and familial forms of PD/DLB, and the assay parameters show poor correlations with various clinical measures. End-point dilution analysis in αSyn RT-QuIC assays allows for the quantitation of relative amounts of αSyn(D) seeding activity that may correlate moderately with clinical measures and levels of other biomarkers. Herein, we review recent advancements in α-synuclein SAAs for detecting αSyn(D) and describe in detail the modified Spearman–Karber quantification algorithm used with end-point dilutions.