Abstract: Synthesis and applications of fluorescent dyes which are derivatives of benzo[c]xanthenes is described. The dyes exhibit pH dependent absorption and fluorescence spectra with pKas near the normal physiological range. Unlike fluorescein, the dyes exhibit emission of different characteristic wavelengths dependent on the pH of the medium. This permits several methods of measuring the pH of the medium in contact with the indicator including measuring two emissions with one excitation, selectively exiting the acid and base forms independently and measuring their emission at either single or dual wavelengths, or measuring the characteristic pH dependent absorption or fluorescence excitation spectral. Methods are presented for making the indicators permeant to cell membranes for the measurement of intracellular pH.