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An ESR, Mass Spectrometry and Fluorescence Microscopy Approach to Study the Stearic Acid Derivatives Anchoring in Cells

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Abstract

Lateral phase separations in biological membranes are of great interest, making electron spin resonance (ESR) spectroscopy combined with spin labeling a nondestructive and sensitive technique for the study of lipid rafts. It is currently accepted that the spin probe is localized on the plasma membrane. However, no study confirms this hypothesis. Herein, we report, for the first time, an accurate multispectral method for the quantification of the lipid spin label presence in every subcellular fraction. Cells were incubated with the 5-DOXYL stearic acid derivative and then subfractionated. Results of our multimodal spectroscopy approach ubiquitously demonstrate that the ESR spin label presence only sets in the plasma membranes.

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Acknowledgments

We offer special thanks to Dr. Mazzucchelli, from the Laboratory of Mass Spectrometry (GIGA-R and CART, University of Liège, Belgium), for his advice in terms of expert knowledge in the field of cell subfractionation. We also thank Dr. DePauw-Gillet, from the Laboratory of Histology–Cytology (University of Liège, Belgium) for her help and advice on cell extraction. The authors thank Dr. Mouithys-Mickalad for his scientific advice. We offer special thanks to Mr. Cobraiville for his help in the achievement of extractions. M.C. benefited from a FRIA (FNRS) PhD fellowship. M.F. is a research associate at F.R.S.-FNRS (National Fund for Scientific Research). We thank the FNRS for its financial support.

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Authors and Affiliations

  1. Laboratory of Biomedical Spectroscopy, Department of Physics, B5, University of Liège, Sart-Tilman, 4000, Liège, Belgium

    A. Grammenos, C. Quoilin & M. Hoebeke

  2. Laboratory of Analytical Pharmaceutical Chemistry, Department of Pharmacy, Drug Research Center, University of Liège, 1 av. de l’Hôpital, 4000, Liège, Belgium

    M. Fillet

  3. Laboratory of Histology-Cytology, Departement of Chemistry, University of Liège, Sart-Tilman, 4000, Liège, Belgium

    M. Collodoro

  4. Laboratory of Mass Spectrometry, GIGA-R and CART, Department of Chemistry, University of Liège, Sart-Tilman, 4000, Liège, Belgium

    M. Collodoro

  5. Center of Biomedical Photonic of LUMAT Federation, University of Paris-Sud, Orsay, France

    S. Lecart

  6. Institut of Molecular Sciences of Orsay, University of Paris-Sud, Orsay, France

    M.-P. Fontaine-Aupart

Authors
  1. A. Grammenos
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  2. M. Fillet
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  3. M. Collodoro
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  4. S. Lecart
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  5. C. Quoilin
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  6. M.-P. Fontaine-Aupart
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  7. M. Hoebeke
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Correspondence to A. Grammenos.

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Grammenos, A., Fillet, M., Collodoro, M. et al. An ESR, Mass Spectrometry and Fluorescence Microscopy Approach to Study the Stearic Acid Derivatives Anchoring in Cells. Appl Magn Reson 43, 311–320 (2012). https://doi.org/10.1007/s00723-012-0375-0

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  • DOI: https://doi.org/10.1007/s00723-012-0375-0

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