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of Molecular Medicine and bByrd Alzheimer's Research Institute, Morsani College of Medicine, University of South Florida, Tampa, FL
Background
Methods
Results
Amount
Function
Tau (383)
16 M
Protein
5-150 nM
Experimental Variable
Thioflavin T
5 M
Fluorescence
EDTA
10 mM
Monomer
Dimer
Among three experimental antibodies A10, 181 and 396, only antibody
396 produced a dramatic increase in the nucleation rate constant,
corresponding to a shortened lag phase across all concentrations of
antibody. While addition of 396 significantly altered the nucleation rate
constant, increases in concentration of the antibody showed no
discernible influence.
NaCl
100 mM
Hepes, pH 7.5
10 mM
pH Buffer
Heparin Sulfate
5 M
Facilitate Aggregation
Protofibrils
Mature Fibrils
Nucleation
Antibody Binding Sites on Tau
Elongation/Fibrillation
Time (Hours)
Fluorescence
Fluorescence
Data Fitting
Electron Microscopy
Yo
Xo
Four Parameter Sigmoid
Curve
Time (Hours)
Objectives
The mechanism of antibody-mediated Tau therapy remains
elusive. In-vitro kinetic aggregation assays containing very low
concentrations of antibodies may help facilitate the
understanding of antibody-Tau interactions at physiological
levels. We aim to examine the kinetic effect of three antibodies
(A10, 181, and 396) on the formation of Tau fibrils.
Control
35 nM,
Ab 396
Control
65 nM,
Ab 396
Control
110 nM,
Ab 396
Conclusions
We observed that addition of the antibody 396 to the microtubule
associated protein Tau in the process of its aggregation resulted
in an increase in the rate of lag phase nucleation paralleled by
an increase in the rate of active elongation/fibrillation.
Although aggregation rates had undergone a sizable increase,
electron microscopy revealed retention of oligomers and a
reduction in the density of end product fibrils. Perhaps these
states represent a less toxic form of Tau inclusions that form in
the presence of the antibody.