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Validation of TPEN as a Zinc Chelator in Fluorescence Probing of Calcium in Cells with the Indicator Fura-2

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Abstract

Fura-2 is widely used as a fluorescent probe to monitor dynamic changes in cytosolic free calcium in cells, where Ca2+ can enter through several types of voltage-operated or ligand-gated channels. However, Fura-2 is also sensitive to other metal ions, such as zinc, which may be involved in ionic channels and receptors. There is interest, in particular, in studying the synapses between mossy fibers and CA3 pyramidal cells which contain both calcium and high quantities of free or loosely bound zinc. We have found, through fluorescence probing, that endogenous zinc inhibits mossy fiber calcium transients. However, since these results might be explained by an effect of the zinc chelator N,N,N’,N’-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) on the spectral properties of Fura-2, we have carried out a validation of the method through fluorescence excitation spectra of the complex Fura-2/calcium, and show that TPEN does not affect these spectra. This supports the idea that the observed calcium enhancement is related to a zinc inhibition of presynaptic calcium mechanisms, and confirms the use of the chelator TPEN as a general procedure for the biophysical study of Ca(II) in the presence of Zn(II) using Fura-2.

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Authors and Affiliations

  1. Department of Physics, University of Trás-os-Montes and Alto Douro (UTAD), 5000-911, Vila Real, Portugal

    Carlos M. Matias

  2. Department of Chemistry, University of Beira Interior, 6201-901, Covilhã, Portugal

    João M. Sousa & Mona Arif

  3. Department of Physics and Center for Neurosciences of Coimbra, University of Coimbra, 3004-516, Coimbra, Portugal

    M. Emília Quinta-Ferreira

  4. Departamento de Química, Universidade de Coimbra, 3004-535, Coimbra, Portugal

    Hugh D. Burrows

Authors
  1. Carlos M. Matias
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  2. João M. Sousa
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  3. M. Emília Quinta-Ferreira
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  4. Mona Arif
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  5. Hugh D. Burrows
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Correspondence to M. Emília Quinta-Ferreira or Hugh D. Burrows.

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Matias, C.M., Sousa, J.M., Quinta-Ferreira, M.E. et al. Validation of TPEN as a Zinc Chelator in Fluorescence Probing of Calcium in Cells with the Indicator Fura-2. J Fluoresc 20, 377–380 (2010). https://doi.org/10.1007/s10895-009-0539-y

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  • DOI: https://doi.org/10.1007/s10895-009-0539-y

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