Monitoring the earliest amyloid-β oligomers via quantized photobleaching of dye-labeled peptides

Misfolding and aggregation of amyloid-beta peptide (A beta) are widely recognized as causative events in Alzheimer's disease (AD). Contrary to earlier hypotheses, recent studies have identified soluble A beta oligomers as the pathogenic agents and documented neurodegenerative effects from species as small as dimers and trimers. As such, detection and characterization of the earliest A beta oligomers are paramount to understanding, preventing, and treating AD. We have exploited quantized photobleaching from individual dye-labeled A beta peptides to characterize monomers and small oligomers tethered to the surface of functionalized coverslips. In this way, we have directly determined reproducible distributions of peptide species in various sample environments. Fresh samples (30 pM peptide) at pH 7.4 consist primarily of monomers and dimers. Both acidic conditions (pH 5.8) and zinc coordination promote greater association, which is further increased in samples prepared from more concentrated stocks. Acid-or zinc-promoted association is largely prevented and/or reversed by addition of the beta-sheet breaker peptide iA beta 5 or the chelator clioquinol, respectively. These results are qualitatively consistent with bulk-solution studies of A beta 40 and demonstrate a powerful approach for definitively characterizing A beta oligomers. The methodology utilized here holds promise for assessing aggregation promoters and inhibitors and investigating peptide association in its earliest stages. (C) 2008 Elsevier Inc. All rights reserved.