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BioAssay: AID 624121

SAR analysis for the identification of selective inhibitors of the two-pore domain potassium channel (KCNK9): Manual Electrophysiology

The purpose of this assay is to evaluate compound inhibition of KCNK9 two pore domain potassium channels using conventional voltage clamp technique. Whole cell recordings were made from HEK293 cells stably expressing KCNK9 channels. ..more
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 Tested Compounds
 Tested Compounds
All(1)
 
 
Active(1)
 
 
 Tested Substances
 Tested Substances
All(1)
 
 
Active(1)
 
 
AID: 624121
Data Source: Johns Hopkins Ion Channel Center (JHICC_KCNK9_inh_Manual EP)
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2012-05-04
Hold-until Date: 2013-05-01
Modify Date: 2013-05-02

Data Table ( Complete ):           View Active Data    View All Data
Target
BioActive Compound: 1
Related Experiments
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AIDNameTypeComment
488964Summary of probe development for inhibitors of the two-pore domain potassium channel KCNK9Summarydepositor-specified cross reference
488922Primary cell-based screen for identification of compounds that inhibit the two-pore domain potassium channel KCNK9Screeningsame project related to Summary assay
492992Confirmatory screen for identification of compounds that inhibit the two-pore domain potassium channel (KCNK9)Screeningsame project related to Summary assay
492993Specificity screen against Kir2.1 for compounds that modulate the two-pore domain potassium channel (KCNK9)Screeningsame project related to Summary assay
492997Second counter screen for compounds that modulate the two-pore domain potassium channel (KCNK9)Screeningsame project related to Summary assay
504846SAR analysis for compounds that inhibit the two-pore domain potassium channel KCNK9Confirmatorysame project related to Summary assay
504902SAR Analysis for the identification of selective inhibitors of KCNK9 against parental cells: FluxOR Assay CRCConfirmatorysame project related to Summary assay
504920SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in hERG expressing cells: FluxOR Assay CRC for hERG SpecificityConfirmatorysame project related to Summary assay
504922SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in Kir2.1 expressing cells: FluxOR Assay CRCConfirmatorysame project related to Summary assay
540321SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in Kir2.1 expressing cells: FluxOR Assay CRC 2Confirmatorysame project related to Summary assay
540322SAR analysis for compounds that inhibit the two-pore domain potassium channel KCNK9 IIConfirmatorysame project related to Summary assay
540323SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in hERG expressing cells: FluxOR Assay CRC 2Confirmatorysame project related to Summary assay
540324SAR Analysis for the identification of selective inhibitors of KCNK9 against parental cells: FluxOR Assay CRC 2Confirmatorysame project related to Summary assay
588724SAR analysis for compounds that inhibit the two-pore domain potassium channel KCNK9 IIIConfirmatorysame project related to Summary assay
588741SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in hERG expressing cells: FluxOR Assay CRC 3Confirmatorysame project related to Summary assay
588759SAR Analysis for the identification of selective inhibitors of KCNK9 against parental cells: FluxOR Assay CRC 3Confirmatorysame project related to Summary assay
588760SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in Kir2.1 expressing cells: FluxOR Assay CRC 3Confirmatorysame project related to Summary assay
588761SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in KCNQ2 expressing cells: FluxOR Assay CRCConfirmatorysame project related to Summary assay
588776SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in KCNK3 expressing cells: FluxOR Assay CRC 1Confirmatorysame project related to Summary assay
588798SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in KCNQ2 expressing cells: FluxOR Assay CRC 3Confirmatorysame project related to Summary assay
588800SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in KCNQ2 expressing cells: FluxOR Assay CRC 2Confirmatorysame project related to Summary assay
623881SAR Analysis for the identification of selective inhibitors of the two-pore domain potassium channel KCNK9 in Kir2.1 expressing cells: FluxOR Assay CRC 4Confirmatorysame project related to Summary assay
623883SAR Analysis for the identification of inhibitors of the two-pore domain potassium channel KCNK9 - Selectivity assay against hERG: FluxOR Assay CRC 4Confirmatorysame project related to Summary assay
623884SAR Analysis for the identification of selective inhibitors of KCNK9 against parental cells: FluxOR Assay CRC 4Confirmatorysame project related to Summary assay
623886SAR Analysis for the identification of inhibitors of the two-pore domain potassium channel KCNK9 - Selectivity assay against KCNQ2: FluxOR Assay CRCConfirmatorysame project related to Summary assay
623887SAR analysis for compounds that inhibit the two-pore domain potassium channel KCNK9 4Confirmatorysame project related to Summary assay
623894SAR Analysis for the identification of inhibitors of the two-pore domain potassium channel KCNK9 - Selectivity assay against KCNK3: FluxOR Assay CRC 3Confirmatorysame project related to Summary assay
623898SAR analysis for the identification of selective inhibitors of the two-pore domain potassium channel (KCNK9): Automated ElectrophysiologyConfirmatorysame project related to Summary assay
623912SAR Analysis for the identification of inhibitors of the two-pore domain potassium channel KCNK9 - Selectivity assay against KCNK3: FluxOR Assay CRC 2Confirmatorysame project related to Summary assay
623913SAR Analysis for the identification of inhibitors of the two-pore domain potassium channel KCNK9 - Selectivity assay against KCNQ2: FluxOR Assay CRC 4Confirmatorysame project related to Summary assay
623914SAR Analysis for the identification of inhibitors of the two-pore domain potassium channel KCNK9 - Selectivity assay against KCNQ2: FluxOR Assay CRC 2Confirmatorysame project related to Summary assay
623915SAR analysis for the identification of selective inhibitors of the two-pore domain potassium channel (KCNK9) -selectivity assay against KCNK3: Automated ElectrophysiologyConfirmatorysame project related to Summary assay
Description:
Principle of the assay
The purpose of this assay is to evaluate compound inhibition of KCNK9 two pore domain potassium channels using conventional voltage clamp technique. Whole cell recordings were made from HEK293 cells stably expressing KCNK9 channels.
Protocol
1. Cell culture: Cells are routinely cultured in DMEM/high glucose medium, supplemented with 10% Fetal Bovine Serum (HiFBS), 15 microg/mL Blasticidin S and 400 microg/mL hygromycin
2. The stable HEK 293 cell line expressing inducible KCNK9 expression was seeded on to poly-D-lysine coated cover slips two days before testing and induced with 1microg/ml tetracycline 16-24h prior to recording.
3. Recording pipettes are pulled with borosilicate glass (World Precision Instruments Inc, Sarasota, FL) to 2-5 M ohm when filled with a pipette solution containing 140 mM KCl, 1 mM MgCl2, 5 mM EGTA, 5 mM HEPES, 1 mM MgATP, pH 7.3 and placed in the bath solution containing 140 mM NaCl, 5 mM KCl, 0.5 mM CaCl2,1 mM MgCl2, and 10 mM HEPES, pH adjusted to 7.3 with NaOH. Isolated cells are voltage-clamped in whole-cell mode with an AxonPatch 200B amplifier (Axon Instruments Inc, Union City, CA), and currents recorded at 5 kHz. KCNK9 currents were measured by voltage ramps from -120 mV to 40 mV applied every 30 seconds. The patched cells are continuously perfused with bath solutions through a gravity-driven perfusion system.
4. Percent inhibition of KCNK9 currents was calculated as 100x (I5k-Itest)/(I5k) where I is the measured current at -20 mV.
5. Outcome assignment
If the test compound causes greater than 50% inhibition of KCNK9 at 1 microM, the compound is considered to be active (Outcome=2). Otherwise, the compound is designated as inactive (Outcome=1).
6. Score assignment
An inactive test compound is assigned the score of 0. An active test compound is assigned a score between 1 and 100 according to the following criteria: 25 for compounds with 07. Result Definitions:
A. Percent inhibition at 1 microM.
B. Standard deviation of percent inhibition values.
C. Number of cells included in each determination.
Categorized Comment - additional comments and annotations
From PubChem:
Assay Cell Type: HEK293
Result Definitions
TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1Inhibition (%) (1μM**)Percent inhibition of current at 1 uMFloat
2Standard deviationStandard deviation of percent inhibitionFloat
3Number of cellsNumber of cells testedInteger

** Test Concentration.
Additional Information
Grant Number: R03 MH090849-01

Data Table (Concise)
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