Heterologous expression of KCNQ2 (Kv7.2) results in the formation of a slowly activating, noninactivating, voltage-gated potassium channel. Using a cell line that stably expresses KCNQ2, we developed a rubidium flux assay to measure the functional activity and pharmacological modulation of this ion channel. Rubidium flux was performed in a 96-well microtiter plate format; rubidium was quantified using an automated atomic absorption spectrometer to enable screening of 1000 data points/day. Cells accumulated rubidium at 37 °C in a monoexponential manner with t 1/2=40 min. Treating cells with elevated extracellular potassium caused membrane depolarization and stimulation of rubidium efflux through KCNQ2. The rate of rubidium efflux increased with increasing extracellular potassium: the t 1/2 at 50 mM potassium was 5.1 min. Potassium-stimulated efflux was potentiated by the anticonvulsant drug retigabine (EC 50=0.5 μM). Both potassium-induced and retigabine-facilitated efflux were blocked by TEA (IC 50s=0.4 and 0.3 mM, respectively) and the neurotransmitter release enhancers and putative cognition enhancers linopirdine (IC 50s=2.3 and 7.1 μM, respectively) and XE991 (IC 50s=0.3 and 0.9 μM, respectively). Screening a collection of ion channel modulators revealed additional inhibitors including clofilium (IC 50 = 27 μM). These studies extend the pharmacological profile of KCNQ2 and demonstrate the feasibility of using this assay system to rapidly screen for compounds that modulate the function of KCNQ2.