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BioAssay: AID 1720

Sedimentation Assay for Inhibitors of Tau Fibrillization

The microtubule-associated protein tau is an abundant protein in the axons of neurons that stabilizes microtubules. With its ability to modulate microtubule dynamics, tau contributes directly or indirectly, to key structural and regulatory cellular functions. Particularly important is the influence tau exerts on axonal transport, which allows signaling molecules, trophic factors and other more ..
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 Tested Compounds
 Tested Compounds
All(104)
 
 
Active(45)
 
 
Inactive(42)
 
 
Inconclusive(17)
 
 
 Tested Substances
 Tested Substances
All(107)
 
 
Active(48)
 
 
Inactive(42)
 
 
Inconclusive(17)
 
 
AID: 1720
Data Source: NCGC (TAU2734)
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2009-05-06

Data Table ( Complete ):           Active    All
Target
BioActive Compounds: 45
Depositor Specified Assays
AIDNameTypeComment
1460qHTS for Inhibitors of Tau Fibril Formation, Thioflavin T Bindingconfirmatory
1468qHTS for Inhibitors of Tau Fibril Formation, Fluorescence Polarizationconfirmatory
1558Confirmation Concentration-Response Assay for for Inhibitors of Tau Fibril Formation, Thioflavin T Bindingconfirmatory
1475Quantitative High-Throughput Screen for Inhibitors of Tau Fibril Formation: Summarysummary
Description:
NIH Molecular Libraries Probe Production Network [MLPCN]
NIH Chemical Genomics Center [NCGC]

MLPCN Grant: X01 MH083262-01
Assay Provider: Carlo Ballatore, University of Pennsylvania


NCGC Assay Overview:
The microtubule-associated protein tau is an abundant protein in the axons of neurons that stabilizes microtubules. With its ability to modulate microtubule dynamics, tau contributes directly or indirectly, to key structural and regulatory cellular functions. Particularly important is the influence tau exerts on axonal transport, which allows signaling molecules, trophic factors and other essential cellular constituents to travel along the axons. Under pathological conditions, tau becomes sequestered into insoluble aggregates called neurofibrillary tangles. This phenomenon is believed to have pathological consequences by promoting axonal transport deficits that ultimately lead to synaptic dysfunction and neuronal loss. The ability of compounds to block tau fibrillization was assessed using a sedimentation assay. Unlike tau monomers, tau fibrils readily sediment upon centrifugation. Samples from the tau ThT confirmation assay treated with 100 uM compound (AID 1558) were centrifuged and the amount of tau present in the supernatant and pellet was determined.

Crowe, A., Ballatore, C., Hyde, E., Trojanowski, J. Q., and Lee, V. M. Y. (2007) Biochemical and Biophysical Research Communications 358, 1-6.
Protocol
NCGC Assay Protocol Summary:
Human K18 P301L tau (15 uM) in assay buffer (20 uM heparin, 12.5 uM Thioflavine T, 100 mM sodium acetate pH 7) was centrifuged at 186,000 x g for 30 min and the supernatant removed from the pellet. Pellets were resuspended in a volume equal to the supernatant and equal amounts of supernatants and pellet were analyzed by SDS-PAGE using a 12.5% acrylamide gel. The gel was stained with Coomassie Blue and the percentage of tau in each fraction was determined by densitometry.
Comment
Compound Ranking:

Compounds are scored based on their percentage of tau fibrillization inhibition. The actual % inhibition values are used as scores for active compounds. Inactive compounds are assigned a score of 0. Inconclusive compounds are assigned a score of 20.
Result Definitions
TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1PhenotypeIndicates type of activity observed: inhibitor, activator, fluorescent, cytotoxic, inactive, or inconclusive.String
2Inhibition (%)Percentage of tau fibrillization inhibition observed for compound.Float%
3Compound QCNCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling'String
Additional Information
Grant Number: X01 MH083262-01

Data Table (Concise)
Classification
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