SensoLyte« ThT A▀40 and A▀42 Aggregation kits, using the fluorescent dye Thioflavin
T (ThT), provide a convenient and standard method to measure A▀40 and A▀42 aggregation,
respectively. In each of the kits, A▀40 or A▀42 peptide is pretreated to ensure
it is in a monomeric state. Included in the kits is an optimized fibrillation
buffer and two known inhibitors as controls. The assay is based on the property
of ThT dye in which fluorescence (Ex/Em=440/484 nm) increase is proportional
to the amount of dye bound to aggregated A▀ peptides.
Alzheimer's disease (AD), the most common cause of dementia, is characterized
by the presence of senile plaques and neurofibrillary tangles, surrounded by
damaged neurons. Beta-Amyloid (A▀) peptides, A▀40 (1-40) and A▀42 (1-42), were
found to be a major component of the above plaques. Many studies suggest that
these peptides can form toxic oligomers and fibrils under physiological conditions
and rapidly aggregate. Since A▀ aggregation is evidently an essential event
in the pathogenesis of AD, a reliable assay is important to study A▀ fibrillation
kinetics and for screening A▀ aggregation inhibitors.
Figure 1. Fluorescence signal
is correlated with an increase of A▀40 fibril formation. Phenol Red, Morin, Rhodamine
B, and Tannic Acid were added at 50 ÁM final concentration to inhibit A▀40 aggregation.
Fluorescence was monitored at Ex/Em= 440/484 nm every 5 minutes at 37║C with 15
seconds shaking between reads (Flexstation II-384 fluorimeter, Molecular Devices).
Figure 2. Fluorescence signal
is correlated with an increase of A▀42 fibril formation. Phenol Red, Morin, Tannic
Acid, Dopamine, and o-Vanillin were added at 100ÁM final concentration to inhibit
A▀42 aggregation. Fluorescence was monitored at Ex/Em= 440/484 nm every 5 minutes
at 37║C with 15 seconds shaking between reads (Flexstation II-384 fluorimeter,
Figure 3. Thioflavin T staining
of b-amyloid aggregates in an AD brain Image courtesy of Drs. J-P. Guo and Patrick
L. McGeer (Kinsmen Laboratory of Neurological Research, University of British
Columbia, Vancouver, Canada).
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