| Assay Method Information | |
| | Electrophysiology Assay |
| Description: | A programmable valve-linked pressurized perfusion system was used for local application of compounds nearby the cell recorded in a consistent flow rate of 2-3 ml/min. Series resistance was corrected and data were sampled at 5 kHz and low pass filtered at 2.4 kHz using MultiClamp 700B amplifier with pCLAMP11 software (Molecular Devices, USA). To evaluate the effect of test compounds on hKv7.2/3 currents at −40 mV membrane potential (the threshold for action potential initiation) a −40 mV pulse train protocol was conducted: Membrane potential is held on −90 mV and is then clamped to −40 mV for 1.5 seconds, followed by clamping the membrane to −60 mV to obtain tail currents for 0.75 seconds, and back to the −90 mV holding potential. An interval of 30 seconds in −90 mV holding potential is kept between the sweeps. After a steady baseline current is achieved, test compound is locally applied using the pressurized perfusion system until a maximal and stable channel modulation in −40 mV is achieved, as confirmed by recording of three similar consequent responses. Thereafter, in a similar manner, cells are perfused back to their control bath solution, to assess the reversibility of the effect of the compounds. |
| Affinity data for this assay | |
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