Real-time imaging and quantification of amyloid-beta peptide aggregates by novel quantum-dot nanoprobes.
BACKGROUND
Protein aggregation plays a major role in the pathogenesis of neurodegenerative disorders, such as Alzheimer's disease. However, direct real-time imaging of protein aggregation, including oligomerization and fibrillization, has never been achieved. Here we demonstrate the preparation of fluorescent semiconductor nanocrystal (quantum dot; QD)-labeled amyloid-beta peptide (QDAbeta) and its advanced applications.


RESULTS
The QDAbeta construct retained Abeta oligomer-forming ability, and the sizes of these oligomers could be estimated from the relative fluorescence intensities of the imaged spots. Both QDAbeta coaggregation with intact Abeta42 and insertion into fibrils were detected by fluorescence microscopy. The coaggregation process was observed by real-time 3D imaging using slit-scanning confocal microscopy, which showed a typical sigmoid curve with 1.5 h in the lag-time and 12 h until saturation. Inhibition of coaggregation using an anti-Abeta antibody can be observed as 3D images on a microscopic scale. Microglia ingested monomeric QDAbeta more significantly than oligomeric QDAbeta, and the ingested QDAbeta was mainly accumulated in the lysosome.


CONCLUSIONS
These data demonstrate that QDAbeta is a novel nanoprobe for studying Abeta oligomerization and fibrillization in multiple modalities and may be applicable for high-throughput drug screening systems.
Tokuraku
Kiyotaka
K
Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, Nebraska, United States of America.
Marquardt
Meg
M
Ikezu
Tsuneya
T
D000682:Amyloid / Q000648:ultrastructure; D016229:Amyloid beta-Peptides / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D002470:Cell Survival; D002478:Cells, Cultured; D006801:Humans; D051379:Mice; D017628:Microglia / Q000166:cytology / Q000378:metabolism; D057054:Molecular Imaging / Q000379:methods*; D053758:Nanoparticles / Q000737:chemistry* / Q000648:ultrastructure; D020836:Protein Structure, Quaternary; D045663:Quantum Dots; D013997:Time Factors
D000682:Amyloid; D016229:Amyloid beta-Peptides