Advances in fluorescence spectroscopy: Multi-photon excitation, engineeredproteins, modulation sensing and microsecond rhenium metal-ligand complexes

The technology and applications of fluorescence spectroscopy are rapidly ad vancing. In this overview presentation we summarize some recent development s from this laboratory. Two and three-photon excitation have been observed for a wide variety of intrinsic and extrinsic fluorophores, including trypt ophan, tyrosine, DNA stains, membrane probes, and even alkanes. It has been possible to observe multi-photon excitation of biopolymers without obvious photochemical or photo-thermal effects. Although not described in our lect ure, another area of increasing interest is the use of engineered proteins for chemical and clinical sensing. We show results for the glucose-galactos e binding protein from E. coli. The labeled protein shows spectral changes in response to micromolar concentrations of glucose. This protein was used with a novel sensing method based on the modulated emission of the labeled proteins and a long lifetime reference fluorophore. And finally, we describ e a recently developed rhenium complex which displays a lifetime near 3 mu s in oxygenated aqueous solution. Such long Lifetime probes allow detection of microsecond dynamic processes, bypassing the usual nanosecond timescale limit of fluorescence. The result of these developments in protein enginee ring, sensing methods, and metal-ligand probe chemistry will be the increas ed use of fluorescence in clinical chemistry and point-of-care analyses.