A kinetic aggregation assay allowing selective and sensitive amyloid-? quantification in cells and tissues.

ABSTRACT: The process of amyloid-? (A?) fibril formation is genetically and pathologically linked to Alzheimer's disease (AD). Thus, a selective and sensitive method for quantifying A? fibrils in complex biological samples allows a variety of hypotheses to be tested. Herein, we report the basis for a quantitative in vitro kinetic aggregation assay that detects seeding-competent A? aggregates in mammalian cell culture media, in Caenorhabditis elegans lysate, and in mouse brain homogenate. Sonicated, proteinase K-treated A? fibril-containing tissue homogenates or cell culture media were added to an initially monomeric A?(1-40) reporter peptide to seed an in vitro nucleated aggregation reaction. The reduction in the half-time (t(50)) of the amyloid growth phase is proportional to the quantity of seeding-competent A? aggregates present in the biological sample. An ion-exchange resin amyloid isolation strategy from complex biological samples is demonstrated as an alternative for improving the sensitivity and linearity of the kinetic aggregation assay.

SUBMITTER: Du D 

PROVIDER: S-EPMC3051019 | biostudies-literature | 2011 Mar

REPOSITORIES: biostudies-literature