Intracellular Ca(2+) mobilization is a useful readout to screen for agonists or antagonists
of G-protein -coupled receptors (GPCRs). Here, we describe methods to conduct high-throughput
screening of stably or transiently transfected HTC4 cells expressing the individual
S1P1-5 receptor subtypes. The cells are grown in 96-well plates and loaded with the
cell permeable fluorescent Ca(2+) indicator dye Fura-2-AM. Changes in intracellular
Ca(2+) levels in response to S1P or test compounds are detected using a FlexStation
II scanning fluorometer with integrated fluidics transfer capabilities.
