Free calcium in isolated chick embryo heart cells measured using quin2 and fura-2

Cytosolic free calcium (Cai) was measured using quin2 and fura-2 in isolated chick embryo heart cells. Account was taken of extracellular quin2 and fura-2 (which could not be entirely washed away) by adding Mn. Shortly after loading with quin2, Cai was 49 nM (n = 7) but then rose continuously at a rate varying between 13 and 88%/h. By varying the time between cell isolation and quin2 loading, it was ascertained that the loading was causing the rise in Cai. In one set of experiments, Cai was stable in time and the apparent Cai increased steadily from 55 to 179 nM as dye loading (quin2 or fura-2) was decreased from 1 mM to 5 microM. We conclude that although quin2 and fura-2 are useful for comparing Cai levels and determining whether Cai changes as a result of certain maneuvers, they do not provide an absolute measure of Cai in isolated embryonic heart cells.