Selectivity and sensitivity of fluorescence lifetime-based metal ion biosensing using a carbonic anhydrase transducer

A key performance criterion for metal ion determinations in complex media like serum, cytoplasm of the cell, and sea water is selectivity: the ability to determine the analyte(s) of interest, in the presence of relatively high concentrations of interferents. Cu(II), Zn(II), Cd(II), Co(II), and Ni(II) may be determined by changes they induce in the fluorescence lifetime and intensity of site-specifically labeled fluorescent variants of apocarbonic anhydrase II. Free metal ion concentrations in the picomolar range (for Cu(II) and Zn(II)) and the nanomolar range (for Cd(II), Co(II), and Ni(II)) were determined, based on the affinity of the apoenzyme for these ions. Mg(II) at 50 mM and Ca(II) at 10 mM produced no effect. By the use of different fluorescent labels, transducers were made which responded well to Cu(II), Co(II), and Ni(II), but not to Zn(II) and Cd(II), and vice versa.