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Fluorescence lifetime characterization of magnesium probes: Improvement of Mg2+ dynamic range and sensitivity using phase-modulation fluorometry

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Abstract

We measured the Mg2+-dependent absorption spectra, emission spectra, quantum yields, and intensity decays of most presently available fluorescent magnesium probes. The lifetimes were found to be strongly Mg2+ dependent for Mag-quin-1, Mag-quin-2, magnesium green, and magnesium orange and increased 2- to 10-fold upon binding of Mg2+. The lifetimes of Mag-fura-2, Mag-fura-5, Mag-fura red, and Mag-indo-1 were similar in the presence and absence of Mg2+. Detailed timeresolved measurements were carried out for Mag-quin-2 and magnesium green using phase-modulation fluorometry. Apparent dissociation constants (K d) were determined from the steady-state and time-resolved data. Their values were compared and discussed. Mg2+ sensing is described using phase and modulation data measured at a single modulation frequency. Phase angle and modulation data showed the possibility of obtaining a wider Mg2+-sensitive range than available from intensity measurements. A significant expansion in the Mg2+-sensitive range was found for Mag-quin-2 using excitation wavelengths from 343 to 375 nm, where the apparentK d from the phase angle was found to vary from 0.3 to about 100 mM. Discrimination against Ca2+ was also measured for Mag-quin-2 and magnesium green. Significant phototransformation and/or photode-composition, which affect the sensitivity to Mg2+, were observed for Mag-quin-2 and magnesium green under intense and long illumination.

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Authors and Affiliations

  1. Center for Fluorescence Spectroscopy and Medical Biotechnology Center, Department of Biological Chemistry, University of Maryland School of Medicine, 108 North Greene Street, 21201, Baltimore, MD

    Henryk Szmacinski & Joseph R. Lakowicz

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  1. Henryk Szmacinski
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  2. Joseph R. Lakowicz
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Szmacinski, H., Lakowicz, J.R. Fluorescence lifetime characterization of magnesium probes: Improvement of Mg2+ dynamic range and sensitivity using phase-modulation fluorometry. J Fluoresc 6, 83–95 (1996). https://doi.org/10.1007/BF00732047

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  • DOI: https://doi.org/10.1007/BF00732047

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