Fig 3. Trap and seed kinetic experiment.
3 μM α-syn seeds without and with pre-incubation with α- or γB-crystallin were trapped by filtration in GHP filter membrane plates with a 200 nm cutoff. The trapped fibrils without any pre-treatment were then incubated for 2 h with freshly purified α-syn monomer and newly filtered ((A) Filtrate 1). The same procedure was performed for the trapped fibrils that had been pre-incubated for 2 h with two concentrations 0.2 and 2 mg/ml of either α-crystallin ((B) Filtrate 2 and 3, respectively) or γB-crystallin ((C) Filtrate 4 and 5, respectively). The flow-through was collected in non-binding PEGylated plates supplemented with ThT and monitored in a plate reader under quiescent conditions at 37°C. The figures show the individual aggregation kinetic traces of at least three experimental repeats.