Fig. 1.

Amyloid seeding at pH 4.3 of wild-type TTR by ex vivo ATTR seeds extracted from the explanted heart of an ATTR-D38A patient. (A) Amyloid seeding assay of recombinant wild-type TTR when 30 ng/μL ATTR-D38A ex vivo seeds were added at 0 h and after 22 h of preincubation, as monitored by ThT fluorescence. (B) Electron micrograph of aggregates of wild-type TTR after 24 h (i and iii) or 4 d (ii and iv) of incubation. ATTR seeds were added at 0 h (i and ii). (Scale bars, 100 nm.) (C and D) Amyloid seeding assays monitored by ThT fluorescence. Increasing amounts of ATTR-D38A ex vivo seeds were added at time 0 to recombinant wild-type TTR (C) or D38A TTR (D). (Right Insets) Anti–His-tag immuno-dot blot of insoluble fractions (IF). (E) Protein concentration in the soluble fraction extracted from C and D, measured by BCA protein assay. The dashed line marks the initial protein concentration (0.5 mg/mL). (F) The 280-nm absorbance of insoluble fractions collected from an amyloid seeding assay of recombinant wild-type TTR with and without 30 ng/L sonicated or nonsonicated seeds. n = 3; error bars indicate SD. AU, absorbance units; a.u., arbitrary units; N/A, not applicable. In A, C, and D all replicates are shown.