In situ cryo-electron tomography of beta-amyloid and tau in post-mortem Alzheimer's disease brain

A defining pathological feature of most neurodegenerative diseases is the assembly of proteins into amyloid that form disease-specific structures. In Alzheimers disease (AD) this is characterised by the deposition of amyloid-{beta} (A{beta}) and tau with AD-specific conformations. The in situ structure of amyloid in the human brain is unknown. Here, using cryogenic fluorescence microscopy (cryoFM)-targeted cryo-sectioning, cryo-focused ion beam scanning electron microscopy (cryoFIB-SEM) liftout and cryo-electron tomography (cryoET), we determined the in-tissue structure of {beta}-amyloid and tau pathology in fresh post-mortem AD donor brain. {beta}-amyloid plaques contained a mixture of fibrils and protofilaments arranged in parallel arrays and lattice-like structures, some of which were branched. Extracellular vesicles, extracellular droplets and open lipid bilayer sheets defined non-amyloid constituents of amyloid plaques. In contrast, tau inclusions were characterised by clusters of unbranched filaments. Subtomogram averaging of filaments within each cluster revealed distinct structures including variably twisted paired helical filaments (PHF) and chronic traumatic encephalopathy (CTE)-like tau filaments that were situated [~]1 m apart within two microscopic regions of pathology. Filaments within a cluster were similar to each other, but different between clusters, showing that fibril heterogeneity is spatially organised and influenced by the subcellular tissue environment. The in situ structural approaches outlined here for targeting specific proteins within human donor tissues have applications to a broad range of neurodegenerative diseases.