Fig. 7: Sizing of multiple species within a heterogenous aggregation mixture by smMDS.

a Schematic of an aggregation reaction composed of monomeric α-synuclein and fibrillar species. b Sizing of α-synuclein fibrils in the presence of an excess of monomeric α-synuclein. Continuous scan diffusion profiles (left panel) for pure monomeric α-synuclein (10 nM) (blue), α-synuclein fibrils (10 nM monomer equivalent) (green), and a mixture of α-synuclein fibrils (9 nM monomer equivalent) and monomeric α-synuclein (1 nM) (pink). The right panel is a zoom-in as indicated by a dashed box in the left panel. Bursts correspond to the passing of fibrils through the confocal detection volume. c Step scan measurement of a mixture of α-synuclein fibrils (9 nM monomer equivalent) and monomeric α-synuclein (1 nM). The top panel shows an exemplary fluorescence time trace (1-ms binning) at diffusion profile position 340 µm, as indicated. An intensity threshold was applied to separate signal from fibrils (red) and monomer (purple). The bottom panels show diffusion profiles created from the fibril and monomer signals, respectively. Diffusion profiles are shown as blue lines, experimental fits as orange lines, and errors as green bands. Extracted hydrodynamic radii R_H [with errors] are given as insets. For definitions of errors, please refer to the legend of Fig. 1b. d Comparison of extracted sizes from triplicate step scan measurements. Shown are R_H of species extracted from a mixture of α-synuclein fibrils (9 nM monomer equivalent) and 1 nM monomeric α-synuclein (red and purple, respectively), pure monomeric α-synuclein (blue), 10 nM monomer equivalent of α-synuclein fibrils (green). Data points (mean) were obtained from triplicate measurements. Error bars denote standard deviations. Step scan measurements of pure α-synuclein (10 nM) (panel e) and pure fibrils (10 nM monomer equivalent) (panel f). The top panels show exemplary fluorescence time traces (1-ms binning) at diffusion profile positions 338 µm and 340 µm, respectively. Diffusion profiles are shown as blue lines, experimental fits as orange lines, and errors as green bands. Extracted R_H [with errors] are given as insets. For definitions of errors, please refer to the legend of Fig. 1b. Source data are provided as a Source Data file.