AA protein in experimental murine AA amyloid fibrils: a high resolution ultrastructural and immunohistochemical study comparing aldehyde-fixed and cryofixed tissues.

In a previous study, the fibrils of experimental murine AA amyloid were found to be microfibril-like structures with the AA protein (in the form of 1 nm wide flexible filaments) on their exterior surface. In this study, we have re-examined the AA amyloid fibrils with advanced methods of cryofixation and freeze substitution which are known to retain ultrastructural detail as close as possible to the living state. The observations were compared to those obtained with conventional methods of aldehyde fixation. Cryofixation and freeze substitution confirmed the microfibril-like nature of the inner part of the AA amyloid fibril. The AA protein was present on the exterior surface in the form of 3 nm wide 'helical rods' formed by the tight coiling of the 1 nm wide AA protein flaments. The 'helical rods' were arranged parallel to the axis of the fibril and to one another with a uniform center-to-center distance of 5 nm. This arrangement was fully preserved in amyloid fibrils after cryofixation and freeze substitution, but was present in only some areas of formaldehyde fixed mouse spleen AA amyloid This conformation and orientation of AA protein is likely to be that in its native state, given the ability of these advanced methods of biological preservation to retain structures close to that of the living state. This information shoulld be of considerable value in comparing the structure of amyloid fibrils observed in situ with those isolated from tissue or generated in vitro.