OPTICAL SENSING OF PH AND PCO2 USING PHASE-MODULATION FLUOROMETRY AND RESONANCE ENERGY-TRANSFER

We have developed optical sensors for pH and/or pCO2 based on phase shift and modulation measurements of time-resolved fluorescence energy transfer. pH-insensitive donors and pH-sensitive acceptors were encased in CO2-permeable silicon membranes. The extent of energy transfer depends on the pH-dependent absorption spectra of the acceptor. The measurements were made insensitive to inner-filter effects of the total intensity by using the phase-modulation method to determine the donor decay times. The general applicability of this method was demonstrated using three donor-acceptor pairs, all of which can be excited with an inexpensive green 543-nm He-Ne laser.