This assay is a confirmation study for the qHTS Assay for Tau Filament Binding (PubChem AID 596). Tau monomers form filaments in vitro in the presence of arachidonic acid. 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. ..more
Related BioAssays
Related BioAssays
Target
| Sequence: Microtubule-associated protein tau [Homo sapiens] Description ..   Protein Family: Tau and MAP protein, tubulin-binding repeat Gene:MAPT Related Protein 3D Structures |
BioActive Compounds: 24
Depositor Specified Assays
| AID | Name | Type | Comment |
|---|---|---|---|
| 596 | qHTS Assay for Tau Filament Binding | confirmatory |
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:
This assay is a confirmation study for the qHTS Assay for Tau Filament Binding (PubChem AID 596). Tau monomers form filaments in vitro in the presence of arachidonic acid. 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.
NIH Chemical Genomics Center [NCGC]
MLSCN Grant: 1 X01 MH079825-01
Assay Provider: Jeffrey A. Kuret, Ohio State University
NCGC Assay Overview:
This assay is a confirmation study for the qHTS Assay for Tau Filament Binding (PubChem AID 596). Tau monomers form filaments in vitro in the presence of arachidonic acid. 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.
Protocol
NCGC Assay Protocol Summary
The assay was performed in 96-well round bottom opaque plates (VWR, West Chester, PA) in 100 uL final volume. Recombinant His6-htau40 was diluted to 5 uM in assembly buffer (10 mM HEPES, pH 7.4, 100 mM NaCl, 5 mM dithiothreitol) containing 50 uM arachidonic acid inducer. ThS probe was then added to a final concentration of 10 uM with test compounds assayed at nine 3-fold dilutions beginning at 10 uM final concentration. Each compound was tested at least twice in separate assays. Controls without protein or without compound were included on each plate. After shaking 2 min to promote mixing, plates were covered with self-adhesive aluminum foil and incubated overnight at 37 C. Fluorescence (ex = 440 nm; em = 485 nm) was measured after brief shaking (10 s) in a FlexStation plate reader (Molecular Devices, Sunnyvale, CA) operated at sensitivity 10, automatic PMT.
A portion of these data are reported in Honson NS, Johnson RL, Huang W, Inglese J, Austin CP, and Kuret J. (2007) Neurobiol Dis. 28(3), 251-60.
The assay was performed in 96-well round bottom opaque plates (VWR, West Chester, PA) in 100 uL final volume. Recombinant His6-htau40 was diluted to 5 uM in assembly buffer (10 mM HEPES, pH 7.4, 100 mM NaCl, 5 mM dithiothreitol) containing 50 uM arachidonic acid inducer. ThS probe was then added to a final concentration of 10 uM with test compounds assayed at nine 3-fold dilutions beginning at 10 uM final concentration. Each compound was tested at least twice in separate assays. Controls without protein or without compound were included on each plate. After shaking 2 min to promote mixing, plates were covered with self-adhesive aluminum foil and incubated overnight at 37 C. Fluorescence (ex = 440 nm; em = 485 nm) was measured after brief shaking (10 s) in a FlexStation plate reader (Molecular Devices, Sunnyvale, CA) operated at sensitivity 10, automatic PMT.
A portion of these data are reported in Honson NS, Johnson RL, Huang W, Inglese J, Austin CP, and Kuret J. (2007) Neurobiol Dis. 28(3), 251-60.
Comment
Compound Ranking:
1. Compounds are first classified as being active, fluorescent (artifactual results), inactive, or inconclusive.
2. Active compounds were ranked by efficacy and potency from 99 to 50. Inconclusive outcome compounds were scored based on the phenotype observed: 25 for less conclusive actives, 10 for likely artifactual data, and 0 for inactive.
1. Compounds are first classified as being active, fluorescent (artifactual results), inactive, or inconclusive.
2. Active compounds were ranked by efficacy and potency from 99 to 50. Inconclusive outcome compounds were scored based on the phenotype observed: 25 for less conclusive actives, 10 for likely artifactual data, and 0 for inactive.
Result Definitions
| TID | Name | Description | Histogram | Type | Unit | |
|---|---|---|---|---|---|---|
| Outcome | The BioAssay activity outcome | Outcome | ||||
| Score | The BioAssay activity ranking score |  | Integer | |||
| 1 | Phenotype | Indicates type of activity observed: active, fluorescent, inactive, or inconclusive. | String | |||
| 2 | Potency* | Concentration at which compound exhibits half-maximal efficacy, AC50. Extrapolated AC50s also include the highest efficacy observed and the concentration of compound at which it was observed. | | Float | μM | |
| 3 | Efficacy | Maximal efficacy of compound, reported as a percentage of control. These values are estimated based on fits of the Hill equation to the dose-response curves. | | Float | % | |
| 4 | Compound QC | NCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling' | String |
* Activity Concentration.
Data Table (Concise)
Classification
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