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A photoinduced electron transfer-based nanoprobe as a marker of acidic organelles in mammalian cells

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Abstract

Photoinduced electron transfer (PET)-based molecular probes have been successfully used for the intracellular imaging of the pH of acidic organelles. In this study, we describe the synthesis and characterization of a novel PET-based pH nanoprobe and its biological application for the signaling of acidic organelles in mammalian cells. A fluorescent ligand sensitive to pH via the PET mechanism that incorporates a thiolated moiety was synthesized and used to stabilize gold nanoparticles (2.4 ± 0.6 nm), yielding a PET-based nanoprobe. The PET nanoprobe was unambiguously characterized by transmission electron microscopy, proton nuclear magnetic resonance, Fourier transform infrared, ultraviolet-visible absorption, and steady-state/time-resolved fluorescence spectroscopies which confirmed the functionalization of the gold nanoparticles with the PET-based ligand. Following a classic PET behavior, the fluorescence emission of the PET-based nanoprobe was quenched in alkaline conditions and enhanced in an acidic environment. The PET-based nanoprobe was used for the intracellular imaging of acidic environments within Chinese hamster ovary cells by confocal laser scanning microscopy. The internalization of the nanoparticles by the cells was confirmed by confocal fluorescence images and also by recording the fluorescence emission spectra of the intracellular PET-based nanoprobe from within the cells. Co-localization experiments using a marker of acidic organelles, LysoTracker Red DND-99, and a marker of autophagosomes, GFP-LC3, confirm that the PET-based nanoprobe acts as marker of acidic organelles and autophagosomes within mammalian cells.

A PET based ligand has been used to functionalize gold nanoparticles to develop a pH sensitive nanoprobe. The fluorescence of the nanoprobe, following the PET mechanism, is enhanced in acidic environments and quenched at neutral pH. A combination of spectroscopy and confocal fluorescence microscopy is used for confirmation of the cellular uptake of the nanoprobe by Chinese hamster ovary cells. The PET-based nanoprobe has been used as a marker of acidic organelles and autophagosomes within the CHO cells

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Acknowledgments

The authors acknowledge the School of Chemistry, University of East Anglia for a studentship for M.J.M. F.G. acknowledges the financial support of the Spanish MICINN (project number CTQ2009-09953) and the Fundació Caixa Castelló-Bancaixa (project number P1·1B2012-41). The authors are grateful to Dr. Colin McDonald (School of Chemistry, University of East Anglia) for the assistance with the TEM images.

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Authors and Affiliations

  1. School of Chemistry, University of East Anglia, Norwich, Norfolk, NR4 7TJ, UK

    María J. Marín & David A. Russell

  2. Departamento de Química Inorgánica y Orgánica, Universitat Jaume I, 12071, Castellón de la Plana, Spain

    Francisco Galindo

  3. School of Biological Sciences, University of East Anglia, Norwich, Norfolk, NR4 7TJ, UK

    Paul Thomas

  4. Norwich Medical School, University of East Anglia, Norwich, Norfolk, NR4 7TJ, UK

    Tom Wileman

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  1. María J. Marín
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  2. Francisco Galindo
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  5. David A. Russell
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Correspondence to Francisco Galindo or David A. Russell.

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Published in the topical collection Optical Nanosensing in Cells with guest editor Francesco Baldini.

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Marín, M.J., Galindo, F., Thomas, P. et al. A photoinduced electron transfer-based nanoprobe as a marker of acidic organelles in mammalian cells. Anal Bioanal Chem 405, 6197–6207 (2013). https://doi.org/10.1007/s00216-013-6905-2

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  • DOI: https://doi.org/10.1007/s00216-013-6905-2

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