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BioAssay: AID 596

qHTS Assay for Tau Filament Binding

Tau monomers form filaments in vitro in the presence of anionic detergents or fatty acids. The dye Thioflavine S (ThS) binds to tau filaments and upon binding, increases in fluorescence several fold. Small molecules that displace ThS binding or prevent filament binding are identified by a reduction in ThS fluorescence. This assay used histidine-tagged, recombinant human tau protein containing the more ..
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 Tested Compounds
 Tested Compounds
All(69666)
 
 
Active(1391)
 
 
Inactive(66726)
 
 
Inconclusive(1843)
 
 
 Tested Substances
 Tested Substances
All(70699)
 
 
Active(1413)
 
 
Inactive(67413)
 
 
Inconclusive(1873)
 
 
 Related BioAssays
 Related BioAssays
AID: 596
Data Source: NCGC (TAUP555)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Screening Center Network
BioAssay Version:
Deposit Date: 2007-02-16
Modify Date: 2007-12-20

Data Table ( Complete ):           Active    All
Target
Sequence: Microtubule-associated protein tau [Homo sapiens]
Description ..   
Protein Family: Tau and MAP protein, tubulin-binding repeat

Gene:MAPT     Related Protein 3D Structures     More BioActivity Data..
BioActive Compounds: 1391
Depositor Specified Assays
AIDNameTypeComment
911Tau ThS binding assayconfirmatory
937Alpha-synuclein ThS binding assayconfirmatory
956Beta amyloid ThS binding assayconfirmatory
991Tau Fibrillization assayconfirmatory
1460qHTS for Inhibitors of Tau Fibril Formation, Thioflavin T Bindingconfirmatory
1463Counterscreen qHTS for Inhibitors of Tau Fibril Formation, Fluorescence Polarizationconfirmatory
1468qHTS for Inhibitors of Tau Fibril Formation, Fluorescence Polarizationconfirmatory
Description:
NIH Molecular Libraries Screening Centers Network [MLSCN]
NIH Chemical Genomics Center [NCGC]

MLSCN Grant: 1 X01 MH079825-01
Assay Provider: Jeffrey A. Kuret, Ohio State University


NCGC Assay Overview:
Tau monomers form filaments in vitro in the presence of anionic detergents or fatty acids. The dye Thioflavine S (ThS) binds to tau filaments and upon binding, increases in fluorescence several fold. Small molecules that displace ThS binding or prevent filament binding are identified by a reduction in ThS fluorescence. This assay used histidine-tagged, recombinant human tau protein containing the products of exons 2, 3, and 10 that was provided by Jeff Kuret, Ohio State University. To form filaments, 1 uM tau was incubated with 30 uM sodium octadecylsulfate (ODS) in reagent buffer (10 uM thioflavine S, 5 mM 1,4-dithioerythreitol (DTT), 100 mM sodium chloride, 10 mM HEPES pH 7.4) for 15 hrs at 37 C.
Protocol
NCGC Assay Protocol Summary:
Assay Steps. Two uL/well of 2 uM tau in reagent buffer (10 uM thioflavine S, 5 mM DTT, 100 mM sodium chloride, 10 mM HEPES pH 7.4) was dispensed into solid black 1536-well plates. Twenty-three nL compound was transferred to the assay plate after which 2 uL/well of 60 uM ODS in reagent buffer was dispensed for a final assay volume of 4 uL /well. Following 15 hours of incubation at 37 C, fluorescence was detected by an Envision (Perkin Elmer) using 450/8 nm excitation and 510/23 nm emission.

Keywords: NIH Roadmap, MLSCN, MLI, MLSMR, qHTS, NCGC, Alzheimer's disease, Neurofibrillary lesions, tau, tubulin assembly, microtubule stability, cytoskeletal integrity, fibrillization, thioflavin dyes, Amyloidgenic
Comment
Compound Ranking:

1. Compounds are first classified as having full titration curves, partial modulation, partial curve (weaker actives), single point activity (at highest concentration only), or inactive. See data field "Curve Classification". For this assay, apparent inhibitors are ranked higher than compounds that showed apparent activation.

2. For all inactive compounds, PUBCHEM_ACTIVITY_SCORE is 0. For all active compounds, a score range was given for each curve class type given above. LogAC50 was used for determining relative score and was scaled to each curve class' score range.
Result Definitions
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TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1Activity DirectionIndicates direction of observed activity: inactive, decreasing, increasing.String
2Curve DescriptionA description of dose-response curve quality. A complete curve has two observed asymptotes; a partial curve may not have attained its second asymptote at the highest concentration tested. High efficacy curves exhibit efficacy greater than 80% of control. Partial efficacies are statistically signficant, but below 80% of control.String
3Log of AC50Log of AC50.Float
4Hill CoefficientHill slope of fitted curve.Float
5Curve R2R^2 fit value of curve. Closer to 1.0 equates to better Hill equation fit.Float
6Curve Infinite ActivityHill Equation Infinite ActivityFloat
7Curve Zero ActivityHill Equation Zero ActivityFloat
8Curve Excluded PointsWhich dose-response titration points were excluded from analysis based on outlier analysis. Each number represents whether a titration point was (1) or was not (0) excluded, for the titration series going from smallest to highest compound concentrations.String
9Curve ClassNumerical encoding of curve description for the fitted Hill equation.Float
10Activity at 0.732nM% Activity at given concentration.Float%
11Activity at 1.636nM% Activity at given concentration.Float%
12Activity at 3.658nM% Activity at given concentration.Float%
13Activity at 8.18nM% Activity at given concentration.Float%
14Activity at 18.291nM% Activity at given concentration.Float%
15Activity at 0.041uM% Activity at given concentration.Float%
16Activity at 0.091uM% Activity at given concentration.Float%
17Activity at 0.204uM% Activity at given concentration.Float%
18Activity at 0.457uM% Activity at given concentration.Float%
19Activity at 1.022uM% Activity at given concentration.Float%
20Activity at 2.286uM% Activity at given concentration.Float%
21Activity at 5.111uM% Activity at given concentration.Float%
22Activity at 11.429uM% Activity at given concentration.Float%
23Activity at 25.556uM% Activity at given concentration.Float%
24Activity at 57.143uM% Activity at given concentration.Float%
25Compound TypeNCGC designation for compound stage: 'qHTS ECL (Exploratory)', 'NIHSMR (MLSCN)', 'Compound Followup', 'Compound Verification', 'Probe Optimization'String

Data Table (Concise)
Classification
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