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BioAssay: AID 773

Counter Screen for Luciferase-based Assay Positives

This functional assay was developed for detection of compounds inhibiting luciferase. These compounds would be observed as false positives of assays employing luciferase-based detection. ..more
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 Tested Compounds
 Tested Compounds
All(270)
 
 
Active(237)
 
 
Inactive(33)
 
 
 Tested Substances
 Tested Substances
All(270)
 
 
Active(237)
 
 
Inactive(33)
 
 
 Related BioAssays
 Related BioAssays
AID: 773
Data Source: Burnham Center for Chemical Genomics (SDCCG-A025-Luciferase)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Screening Center Network
BioAssay Version:
Deposit Date: 2007-07-11
Modify Date: 2010-08-30

Data Table ( Complete ):           View Active Data    View All Data
BioActive Compounds: 237
Related Experiments
AIDNameTypeComment
411qHTS Assay for Inhibitors of Firefly LuciferaseConfirmatorydepositor-specified cross reference
618Luminescent HTS for small molecule inhibitors of MT1-MMP transcriptionConfirmatorydepositor-specified cross reference
620Fluorescent HTS Cytotoxicity/Cell viability assay (HT1080 cells)Confirmatorydepositor-specified cross reference
1006Counter Screen for Luciferase-based Primary Inhibition AssaysScreeningdepositor-specified cross reference
1027Counter Screen for Luciferase-based Primary Stimulation AssaysScreeningdepositor-specified cross reference
Description:
Data Source: Sanford-Burnham Center for Chemical Genomics (SBCCG)
Source Affiliation: Sanford-Burnham Medical Research Institute (SBMRI, San Diego, CA)
Network: NIH Molecular Libraries Screening Centers Network (MLSCN)
Grant Number: XO1 MH78949-01
Assay Provider: Dr. Alex Strongin, Sanford-Burnham Medical Research Institute


This functional assay was developed for detection of compounds inhibiting luciferase. These compounds would be observed as false positives of assays employing luciferase-based detection.

This screening was developed and performed at the Sanford-Burnham Center for Chemical Genomics (SBCCG) as part of the Molecular Library Screening Center Network (MLSCN).
Protocol
1) 4 uL of compounds or control solutions added to 384-well plate:
a) positive control (EDTA, 12.5 mM final concentration) to columns 1-2
b) negative control (10% DMSO) to columns 23-24
c) serially diluted compounds to columns 3-22
2) 8 uL ATP substrate solution (6.4 uM final concentration) and 8 uL ATPLite solution (PerkinElmer) containing luciferase and luciferin were added to the whole plate
3) Luminescence is measured after 10 min at room temperature on an EnVision plate reader (Perkin Elmer).
4) Data analysis was performed using sigmoidal dose-response equation through non-linear regression
Comment
Compounds with EC50 values < 100 uM are defined as 'actives' in the outcome column. Compounds that show <50% inhibition at all the concentration values are defined as 'inactive' in the outcome field. Compounds with EC50 below 195 nM, the lowest tested concentration, were assigned IC50 value of < 0.2 uM and defined as 'active'.

Activity Scoring
Activity scoring rules developed at Sanford-Burnham Center for Chemical Genomics were devised to take into consideration different information available from screening, and in this case include compound potency, the screening stage of the data and compound behavior in the assay. Details of the Scoring System will be published elsewhere.

Briefly, the outline of the scoring system utilized for the Luciferase assay is as follows:
1) First tier (0-40 range) is reserved for primary screening data and therefore is not applicable in this assay.

2) Second tier (41-80 range) is reserved for dose-response confirmation data
a. Inactive compounds are assigned a score value equal 41.
b. For compounds with EC50<100 uM, the score is linearly correlated with a compound#s potency using the following equation

Score = 44 + 6*(pEC50 - 3)*2.6*(exp(-0.5*nH^2)-exp(-1.5*nH^2)),

where pEC50 is negative log(10) of the EC50 value expressed in mole/L concentration units and nH is Hill coefficient value.
c. Compounds with EC50 below the lowest tested concentration are assigned a score value equal 80.

3) Third tier (81-100 range) is reserved for resynthesized confirmed positives and their analogues and is not applicable in this assay.
Categorized Comment - additional comments and annotations
From ChEMBL:
Assay Type: Binding
Assay Cell Type: NULL
Result Definitions
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TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1EC50_QualifierThis qualifier is to be used with the next TID, EC50. If qualifier is "=", EC50 result equals to the value in that column; if qualifier is ">", EC50 result is greater than that value. if qualifier is "<", EC50 result is less than that value.String
2EC50*EC50 value determined using sigmoidal dose response equationFloatμM
3St.Err(EC50)Standard Error of EC50 valueFloatμM
4nHHill coefficient determined using sigmoidal dose response equationFloat
5Mean HighMean luminescence signal of negative controls in the corresponding plateFloatcps
6STD Deviation HighStandard deviation (n=16) of negative controls in the corresponding plateFloatcps
7Mean LowMean luminescence signal of positive controls in the corresponding plateFloatcps
8STD Deviation LowStandard deviation (n=16) of positive controls in the corresponding plateFloatcps

* Activity Concentration.
Additional Information
Grant Number: XO1 MH78949-01

Data Table (Concise)
Data Table ( Complete ):     View Active Data    View All Data
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