Core-shell silver nanoparticles for optical labeling of cells.

Silver nanoparticles have been modified with self-assembled monolayers of hydroxyl-terminated long chain thiols and encapsulated with a silica shell. The resulting core-shell nanoparticles were used as optical labels for cell analysis using flow cytometry and microscopy. The excitation of plasmon resonances in nanoparticles results in strong depolarized scattering of visible light, permitting detection at the single nanoparticle level. The nanoparticles were modified with neutravidin via epoxide-azide coupling chemistry, to which biotinylated antibodies targeting cell surface receptors were bound. The nanoparticle labels exhibited long-term stability in solutions with high salt concentrations without aggregation or silver etching. Labeled cells exhibited two orders of magnitude enhancement of the scattering intensity compared with unlabeled cells.

Probing the efficacy of peptide-based inhibitors against acid- and zinc-promoted oligomerization of amyloid-ß peptide via single-oligomer spectroscopy.

One avenue for prevention and treatment of Alzheimer's disease involves inhibiting the aggregation of amyloid-ß peptide (Aß). Given the deleterious effects reported for Aß dimers and trimers, it is important to investigate inhibition of the earliest association steps. We have employed quantized photobleaching of dye-labeled Aß peptides to characterize four peptide-based inhibitors of fibrillogenesis and/or cytotoxicity, assessing their ability to inhibit association in the smallest oligomers (n=2-5). Inhibitors were tested at acidic pH and in the presence of zinc, conditions that may promote oligomerization in vivo. Distributions of peptide species were constructed by examining dozens of surface-tethered monomers and oligomers, one at a time. Results show that all four inhibitors shift the distribution of Aß species toward monomers; however, efficacies vary for each compound and sample environment. Collectively, these studies highlight promising design strategies for future oligomerization inhibitors, affording insight into oligomer structures and inhibition mechanisms in two physiologically significant environments.

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

Misfolding and aggregation of amyloid-beta peptide (Abeta) are widely recognized as causative events in Alzheimer's disease (AD). Contrary to earlier hypotheses, recent studies have identified soluble Abeta 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 Abeta oligomers are paramount to understanding, preventing, and treating AD. We have exploited quantized photobleaching from individual dye-labeled Abeta 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 (30pM 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 iAbeta5 or the chelator clioquinol, respectively. These results are qualitatively consistent with bulk-solution studies of Abeta40 and demonstrate a powerful approach for definitively characterizing Abeta oligomers. The methodology utilized here holds promise for assessing aggregation promoters and inhibitors and investigating peptide association in its earliest stages.