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. Author manuscript; available in PMC: 2011 Mar 1.
Published in final edited form as: Future Med Chem. 2010 May 1;2(5):757–774. doi: 10.4155/fmc.10.179

Figure 3. Thallium-fux assay of Kir1.1 channel function for high-throughput molecular library screening.

Figure 3

(A) A stable cell line expressing Kir1.1 under the control of a tetracycline-inducible promoter was developed to avoid cytotoxic effects of constitutive Kir1.1 channel expression and subsequent cell line degeneration. Values are means ± SEM current amplitude normalized to cell capacitance and plotted as a function of test voltage in whole-cell patch clamp experiments. Cells were cultured overnight in the absence (control; darker circles) or presence of tetracycline (lighter circles). (B) Fluorescence assay of Kir1.1 channel activity in a 384-well plate using the thallium-sensitive fluorescent dye FluoZin-2. Cells were cultured overnight in the absence (− Tet) or presence (+ Tet) of tetracycline. One well was pretreated with the Kir1.1 channel inhibitor peptide Tertiapin-Q (+ TPNQ) to block channel activity. Addition of extracellular thallium (at 500 s) evoked an abrupt increase in FluoZin-2 fluorescence that was dependent on Kir1.1 channel expression. This assay was used to screen approximately 225,000 small molecules for novel modulators of Kir1.1.

Reproduced with permission from [47].