Dual lifetime referencing as applied to a chloride optical sensor

A membrane with an optical response to chloride has been developed that contains two luminophores that display two largely different decay times. The first luminophore (the "reference") is a chloride-insensitive ruthenium metal-ligand complex possessing a decay time in the microsecond range. The second luminophore is the short-lived chloride-quenchable fluorescent probe lucigenin. Both are contained in a hydrogel matrix and are excited by a blue LED emitting sinusoidally modulated light. Under these conditions, the chloride-dependent fluorescence intensity of lucigenin can be converted in an analyte-dependent fluorescence phase shift that depends on the ratio of the two luminescence intensities and can be measured at modulation frequencies of typically 45 kHz. The dynamic range of this sensor can be adjusted by either varying the ratio of the two luminophores or selecting a particular optical filter combination.