Intracellular calcium ion concentration measurement using a phase-modulation fluorescence lifetime method with compensation for phase shift due to the presence of proteins

The calcium ion concentration in cells was measured by a phase-modulation fluorescence lifetime method with compensation for proteins. A high-accuracy measurement of the calcium ion concentration is best realized by fluorescence lifetime measurements, because the fluorescence lifetime is independent of the fluorescence intensity. The fluorescence intensity is easily varied by the scattering of excitation and emission LIGHT in cells, photobleaching, the concentration of fluorochromes, and wavelength dispersion of optical elements. A phase-modulation fluorescence lifetime measurement, however, provides high accuracy and precision, and can measure not only the calcium ion concentration, but also other ion concentrations, such as that of magnesium, sodium, and potassium. We have examined the phase-modulation fluorescence lifetime shift using protein compensation in cells, and have measured the calcium ion concentration in cells stimulated with bradykinin.