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BioAssay: AID 504909

A Cell Based Secondary Assay To Explore Vero Cell Cytotoxicity of Purified and Synthesized Compounds that Inhibit Mycobacterium Tuberculosis (5)

This functional assay was developed for detection of compounds inhibiting Vero E6 cells viability as a secondary screen to the beta-lactam sensitizing M. tuberculosis bacteriocidal assay. ..more
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 Tested Compounds
 Tested Compounds
All(69)
 
 
Active(15)
 
 
Inactive(54)
 
 
 Tested Substances
 Tested Substances
All(69)
 
 
Active(15)
 
 
Inactive(54)
 
 
 Related BioAssays
 Related BioAssays
AID: 504909
Data Source: Southern Research Specialized Biocontainment Screening Center (TB_Inh_Vero6)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
BioAssay Version:
Deposit Date: 2011-07-05
Hold-until Date: 2012-04-12
Modify Date: 2012-04-12

Data Table ( Complete ):           Active    All
BioActive Compounds: 15
Depositor Specified Assays
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AIDNameTypeComment
449762High Throughput Screening Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in 7H9 Mediaconfirmatoryprimary & confirmatory screen (w/Glycerol)
493198High Throughput Screening Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in 7H9 Media using Purified and Synthesized Compoundsconfirmatoryconfirmatory screen (w/Glycerol)
504556High Throughput Screening Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in 7H9 Media using Purified and Synthesized Compounds (2)confirmatoryconfirmatory screen (w/Glycerol)
504646High Throughput Screening Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in 7H9 Media using Purified and Synthesized Compounds (3)confirmatoryconfirmatory screen (w/Glycerol)
504857High Throughput Screening Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in 7H9 Media using Purified and Synthesized Compounds (4)confirmatoryconfirmatory screen (w/Glycerol)
449764A High Throughput Confirmatory Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in the absence of Glycerolconfirmatorycounter screen (No Glycerol)
493181A High Throughput Confirmatory Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in the absence of Glycerol using Purified and Synthesized Compoundsconfirmatorycounter screen (No Glycerol)
504564A High Throughput Confirmatory Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in the absence of Glycerol using Purified and Synthesized Compounds (2)confirmatorycounter screen (No Glycerol)
504645A High Throughput Confirmatory Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in the absence of Glycerol using Purified and Synthesized Compounds (3)confirmatorycounter screen (No Glycerol)
504860A High Throughput Confirmatory Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in the absence of Glycerol using Purified and Synthesized Compounds (4)confirmatorycounter screen (No Glycerol)
435019A Cell Based Secondary Assay To Explore Cytotoxicity of Compounds that Inhibit Mycobacterium TuberculosisconfirmatoryVero Cytotox
504335A Cell Based Secondary Assay To Explore Cytotoxicity of Purified and Synthesized Compounds that Inhibit Mycobacterium Tuberculosis.confirmatoryVero Cytotox
504562A Cell Based Secondary Assay To Explore Cytotoxicity of Purified and Synthesized Compounds that Inhibit Mycobacterium Tuberculosis (2)confirmatoryVero Cytotox
504684A Cell Based Secondary Assay To Explore Vero Cell Cytotoxicity of Purified and Synthesized Compounds that Inhibit Mycobacterium Tuberculosis (3)confirmatoryVero Cytotox
504854A Cell Based Secondary Assay To Explore Vero Cell Cytotoxicity of Purified and Synthesized Compounds that Inhibit Mycobacterium Tuberculosis (4)confirmatoryVero Cytotox
504642A Cell Based Secondary Assay to Explore Cytotoxicity in HepG2 Cells of Compounds that Modulate Mycobacterium tuberculosis in Media Containing Glycerol versus Media without GlycerolconfirmatoryHepG2 Cytotox
504682A Cell Based Secondary Assay to Explore Cytotoxicity in HepG2 Cells of Compounds that Modulate Mycobacterium tuberculosis in Media Containing Glycerol versus Media without Glycerol (2)confirmatoryHepG2 Cytotox
504853A Cell Based Secondary Assay to Explore Cytotoxicity in HepG2 Cells of Compounds that Modulate Mycobacterium tuberculosis in Media Containing Glycerol versus Media without Glycerol (3)confirmatoryHepG2 Cytotox
504640A Cell Based Secondary Assay to Explore Cytotoxicity in THP-1 Cells of Compounds that Modulate Mycobacterium tuberculosis in Media Containing Glycerol versus Media without GlycerolconfirmatoryTHP1 Cytotox
504683A Cell Based Secondary Assay to Explore Cytotoxicity in THP-1 Cells of Compounds that Modulate Mycobacterium tuberculosis in Media Containing Glycerol versus Media without Glycerol (2)confirmatoryTHP1 Cytotox
504852A Cell Based Secondary Assay to Explore Cytotoxicity in THP-1 Cells of Compounds that Modulate Mycobacterium tuberculosis in Media Containing Glycerol versus Media without Glycerol (3)confirmatoryTHP1 Cytotox
449775Summary of Assays used to Identify Novel Compounds That Inhibit Mycobacterium Tuberculosis in 7H9 Media with GlycerolsummarySummary AID
588437A High Throughput Confirmatory Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in the absence of Glycerol using Purified and Synthesized Compounds (7)confirmatory
588445A Cell Based Secondary Assay To Explore Vero Cell Cytotoxicity of Purified and Synthesized Compounds that Inhibit Mycobacterium Tuberculosis (6)confirmatory
588447High Throughput Screening Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in 7H9 Media using Purified and Synthesized Compounds (8)confirmatory
602431A Cell Based Secondary Assay To Explore Vero Cell Cytotoxicity of Purified and Synthesized Compounds that Inhibit Mycobacterium Tuberculosis (8)confirmatory
602432A Cell Based Secondary Assay to Explore Cytotoxicity in HepG2 Cells of Compounds that Modulate Mycobacterium tuberculosis in Media Containing Glycerol versus Media without Glycerol (6)confirmatory
602433A Cell Based Secondary Assay to Explore Cytotoxicity in THP-1 Cells of Compounds that Modulate Mycobacterium tuberculosis in Media Containing Glycerol versus Media without Glycerol (6)confirmatory
602435High Throughput Screening Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in 7H9 Media using Purified and Synthesized Compounds (9)confirmatory
602437A High Throughput Confirmatory Assay used to Identify Novel Compounds that Inhibit Mycobacterium Tuberculosis in the absence of Glycerol using Purified and Synthesized Compounds (8)confirmatory
Description:
This functional assay was developed for detection of compounds inhibiting Vero E6 cells viability as a secondary screen to the beta-lactam sensitizing M. tuberculosis bacteriocidal assay.

In this assay, we treated Vero E6 cells with compounds selected as hits in the M. tuberculosis assay for 72 hours over a 10 point 2-fold dilution series. Following 72 hours of treatment, relative viable cell number was determined using Cell Titer Glo from Promega. Each plate contained 64 replicates of vehicle treated cells which served as negative controls.
Protocol
Cell Culture: Vero E6 cells were subcultured every 7 days in E-MEM with 10% fetal bovine serum and 2 mM glutamine (complete growth medium), incubated at 37 degrees C in 5% carbon dioxide, and maintained for no more than 20 passages.

Compound Dosing/Plating: Carrier control / compounds were diluted in complete growth medium to prepare a 6X concentrated dosing solution which was dispensed into 384-well black clear-bottom tissue culture treated plates (5 uL volume).

Cell Plating: Twenty uL of complete growth medium containing 3000 cells were dispensed per well. Plates were incubated at 37 C, 5% CO2 for 72h prior to endpoint detection.

Endpoint/Detection: At the end of the treatment period, assay plates were removed from the incubator and equilibrated to room temperature for 10 min. Twenty-five uL of Cell Titer Glo reagent was added and plates were incubated for an additional 10 min in the dark. At the end of the incubation, assay plates were analyzed using a PerkinElmer Envision microplate reader in luminescence mode with an integration time of 0.1 s.

Data Analysis: Sixty-four control wells containing cells treated with DMSO vehicle and were included on each assay plate. Compound data was normalized and reported as % viability which was calculated using the following formula: % viability = 100*(Cmpd Lum-Med background)/(Med Cell Ctrl - Med background). The normalized % viability was plotted against the tested concentrations. The CC50 values were calculated using XLfit formula 205, a 4 parameter Levenburg-Marquardt algorithm with maximum and minimum limits set at 100 and 0, respectively and allowing extrapolation to identify weakly active compounds.
Comment
Outcome: Compounds that showed <70% cell viability for at least one concentration were defined as "Active". If the % viability at all doses was >70%, the compound was defined as "Inactive" and CC50 values were not determined.

Scoring: SRBCSC uses a three-tier scoring system where scores of 0-40 apply to primary screen data, 41-80 indicates confirmatory screen data, and 81-100 is reserved for confirmatory data on resynthesized compounds. Inactive compounds at any screening level receive a score of 0. In this confirmatory screen, "Active" compounds were scored based on Average CC50 results on a tier of 81-100 with "Inactive" compounds scoring 0.
Result Definitions
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TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1Average CC50 ModifierString
2Average CC50*FloatμM
3CC50 Modifier Rep 1String
4CC50 Rep 1FloatμM
5CC50 Std Dev Rep 1Float
6CC50 Hill Slope Rep 1Float
7CC50 NormChi2 Rep 1Float
8CC50 Modifier Rep 2String
9CC50 Rep 2FloatμM
10CC50 Std Dev Rep 2Float
11CC50 Hill Slope Rep 2Float
12CC50 NormChi2 Rep 2Float
13CC50 Modifier Rep 3String
14CC50 Rep 3FloatμM
15CC50 Std Dev Rep 3Float
16CC50 Hill Slope Rep 3Float
17CC50 NormChi2 Rep 3Float
18CC50 Modifier Rep 4String
19CC50 Rep 4FloatμM
20CC50 Std Dev Rep 4Float
21CC50 Hill Slope Rep 4Float
22CC50 NormChi2 Rep 4Float
23Cell Viability @ 100 uM Rep 1 (100μM**)Float%
24Cell Viability @ 50 uM Rep 1 (50μM**)Float%
25Cell Viability @ 25 uM Rep 1 (25μM**)Float%
26Cell Viability @ 12.5 uM Rep 1 (12.5μM**)Float%
27Cell Viability @ 6.25 uM Rep 1 (6.25μM**)Float%
28Cell Viability @ 3.125 uM Rep 1 (3.125μM**)Float%
29Cell Viability @ 1.563 uM Rep 1 (1.563μM**)Float%
30Cell Viability @ 0.781 uM Rep 1 (0.781μM**)Float%
31Cell Viability @ 0.391 uM Rep 1 (0.391μM**)Float%
32Cell Viability @ 0.195 uM Rep 1 (0.195μM**)Float%
33Cell Viability @ 0.098 uM Rep 1 (0.098μM**)Float%
34Cell Viability @ 100 uM Rep 2 (100μM**)Float%
35Cell Viability @ 50 uM Rep 2 (50μM**)Float%
36Cell Viability @ 25 uM Rep 2 (25μM**)Float%
37Cell Viability @ 12.5 uM Rep 2 (12.5μM**)Float%
38Cell Viability @ 6.25 uM Rep 2 (6.25μM**)Float%
39Cell Viability @ 3.125 uM Rep 2 (3.125μM**)Float%
40Cell Viability @ 1.563 uM Rep 2 (1.563μM**)Float%
41Cell Viability @ 0.781 uM Rep 2 (0.781μM**)Float%
42Cell Viability @ 0.391 uM Rep 2 (0.391μM**)Float%
43Cell Viability @ 0.195 uM Rep 2 (0.195μM**)Float%
44Cell Viability @ 100 uM Rep 3 (100μM**)Float%
45Cell Viability @ 50 uM Rep 3 (50μM**)Float%
46Cell Viability @ 25 uM Rep 3 (25μM**)Float%
47Cell Viability @ 12.5 uM Rep 3 (12.5μM**)Float%
48Cell Viability @ 6.25 uM Rep 3 (6.25μM**)Float%
49Cell Viability @ 3.125 uM Rep 3 (3.125μM**)Float%
50Cell Viability @ 1.563 uM Rep 3 (1.563μM**)Float%
51Cell Viability @ 0.781 uM Rep 3 (0.781μM**)Float%
52Cell Viability @ 0.391 uM Rep 3 (0.391μM**)Float%
53Cell Viability @ 0.195 uM Rep 3 (0.195μM**)Float%
54Cell Viability @ 100 uM Rep 4 (100μM**)Float%
55Cell Viability @ 50 uM Rep 4 (50μM**)Float%
56Cell Viability @ 25 uM Rep 4 (25μM**)Float%
57Cell Viability @ 12.5 uM Rep 4 (12.5μM**)Float%
58Cell Viability @ 6.25 uM Rep 4 (6.25μM**)Float%
59Cell Viability @ 3.125 uM Rep 4 (3.125μM**)Float%
60Cell Viability @ 1.563 uM Rep 4 (1.563μM**)Float%
61Cell Viability @ 0.781 uM Rep 4 (0.781μM**)Float%
62Cell Viability @ 0.391 uM Rep 4 (0.391μM**)Float%
63Cell Viability @ 0.195 uM Rep 4 (0.195μM**)Float%
64Cell Viability @ 0.098 uM Rep 4 (0.098μM**)Float%
65VerificationString

* Activity Concentration. ** Test Concentration.
Additional Information
Grant Number: 1 R03 MH084877-01A1

Data Table (Concise)
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