| Assay Method Information | |
| | Forward Mode Assay |
| Description: | The assay is based on the monitoring of intracellular Ca2+concentrations using the PBX Calcium Assay Kit from BD (Becton, Dickinson and Company) with calcium indicator dye 51-9000177BKa (BD, 640177), CHO cells expressing NCX1 were loaded with the dye, and after a preincubation period with the test compound, lonomycin (Calbiochem, 407950) was added. Ionomycin is an ionophor for Ca2+ions mediating an increase of intracellular Ca2+ions. Consequently, intracellular Ca2+ions are exchanged against extracellular Na+ions (Ca2+efflux, forward mode). The decrease of intracellular Ca2+ions was detected by measuring the fluorescence of the calcium indicator dye at a wavelength of 520 nm by a FLIPR device.Briefly, similarly as for the reverse mode, for the forward mode transport assay 18000 cells per well were seeded into a 96 well microplate (Corning COSTAR 3904) and incubated overnight in culture medium (cf. above). A total volume of 100 ul medium per well was used. |
| Affinity data for this assay | |
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