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Assay Method Information

Assay Name:  TREK1 Assay
Description:  The hTREK1 stable cell lines were seeded at a density of 10 000 cells/well in a 12-well plate. Whole-cell membrane currents were amplified using the Axopatch 200A patch-clamp system. Currents were elicited by 1-second voltage ramps descending from +50 mV to -150 mV (from a holding potential of -70 mV). Data acquisition was controlled by pclamp 10.2 software (Molecular Devices, Sunnyvale,CA, USA). A Digidata 1322A interface was used to convert digital-analog signals between amplifier and computer. Data were sampled at 5 kHz and filtered at 2 kHz. Cell membrane capacitance was measured using the 'membrane test' protocolbuilt into pClampex. The bath solution contained (in mM) 150 NaCl, 10 HEPES, 3 KCl, 2 CaCl2, 2 MgCl2, and 5.5 glucose adjusted to pH 7.3 with NaOH. The pipette solution contained (in mM) 145 KCl, 0.5 CaCl2, 10 HEPES, 4 Mg-ATP, 0.3 Na3-GTPadjusted to pH 7.2 with KOH. All experiments were performed at a room temperature of 20-22 °C. TREK1 currents were recorded in the presence of each 10 μm hit compounds. The block percentage of inhibitors was obtained by measuring the percentage of remaining current.
Affinity data for this assay
 

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Last update November 1, 2007
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