To run the assay, cells are loaded with non-fluorescent, thallium-specific FluxOR™ dye (Figure 1). Drugs to be screened are pre-incubated with the cells and the microplates are loaded into the reader, where they are injected with a stimulus buffer containing a low level of thallium ions. The thallium ions freely flow through open potassium channels, acting as a surrogate for K+. When the potassium channel is stimulated, thallium flows into the cell and binds the FluxOR™ dye, generating a fluorescent signal, proportional to channel activity—in physiological saline conditions. Multiple potassium ion channels including Kv1.3 (Figure 2), Kv7.2/Kv7.3 (Figure 3), Kv11.1 (hERG) (Figure 4), Kir1.1, Kir2.1, Kv1.1, Kv2.1 (data not shown) have been demonstrated to work with the FluxOR assay for reliable results.
|FluxOR Potassium Ion Channel Assay||FluxOR II Green Potassium Ion Channel Assay||FluxOR Red Potassium Ion Channel Assay|
|Assay principle||Uses thallium (Tl+ ) ions as surrogate for potassium ions. Thallium is added to the extracellular solution, potassium channels are open, thallium flows down its concentration gradient into the cells, and channel or transporter activity is detected with a proprietary indicator dye that increases in cytosolic fluorescence|
|Performance||Improved indicator dye increases signal to backhground ratio and assay sensitivity||Red emission, can be multiplexed with GFP-expressing or other green fluorescent assays|
|Standard filter set||FITC||RFP|
|Assay protocols||Wash||Wash and No Wash||Wash|
|Avaiable formats||10 microplates||100 microplates||2 microplates||10 microplates||100 microplates||2 microplates||10 microplates|