Table 3.

Comparison of performance between SAA and QSAA

	SAA	QSAA		QSAA in situ
Incubation mode	Sonication/shaking	Quiescent	Quiescent
Cyclic	Cyclic fragmentation	One-step amplification	One-step amplification
Detection device	Fluorescence-plate-reader	Real-time PCR instruments		Incubator
Reaction vessel	96-well plate	96-well PCR plate		24-well plate
Substrate type	Human αSyn (WT/mutant) monomer	Mouse αSyn monomer	Mouse αSyn monomer
Substrate concentration	0.1–1.0 mg/ml	1.0 mg/ml	1.0 mg/ml
AS addition	None	10% w/v AS	10% w/v AS
Beads addition	Silicon/zirconia beads	None	None
Reaction temperature	30–42 ℃	70 ℃	70 ℃
Sample type	Liquid	Liquid		Slices
Incubation duration	24–120 h	6–24 h		6–24 h
Reaction volume	100 μl/well	20 μl/well		200 μl/well
Oil seal	None	Paraffin oil	Paraffin oil
Fluorescent dye	ThT	ThT	ThT
ThT concentration	5 μM	30 μM	30 μM
Quantitative data	Lag phase/Fmax	Lag phase/Fmax		Fluorescence graph
Detection limit (PFFs)	Attograms	Femtograms	Femtograms
Amplification mechanism	Nucleation, elongation	Secondary nucleation, elongation	Secondary nucleation, elongation

SAA, seed aggregation assay; QSAA, quiescent aeed aggregation assay; AS, ammonium sulfate