Ratiometric Ca²⁺ measurements using the FlexStation(®)Scanning Fluorometer

The FlexStation( ® ) Scanning Fluorometer is a fluorescence plate reader that can measure intracellular Ca(2+) concentration using both single-wavelength and dual-wavelength fluorescent probes. The FlexStation uses a Xenon flashlamp and monochromators for both excitation and emission light to allow the use of a wide range of fluorescent indicators. The system incorporates a fluid transfer system for addition of test compounds from a source plate to the cell plate during data acquisition. Both plates are contained within a temperature-controlled unit that can be controlled accurately between room temperature and 45°C. The FlexStation can be configured to read a range of plate sizes. In this chapter generic methods for assessing intracellular Ca(2+) on the FlexStation using ratiometric dyes are described.