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Time resolved and label free monitoring of extracellular metabolites by surface enhanced Raman spectroscopy

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Listed:
  • Victoria Shalabaeva
  • Laura Lovato
  • Rosanna La Rocca
  • Gabriele C Messina
  • Michele Dipalo
  • Ermanno Miele
  • Michela Perrone
  • Francesco Gentile
  • Francesco De Angelis

Abstract

Metabolomics is an emerging field of cell biology that aims at the comprehensive identification of metabolite levels in biological fluids or cells in a specific functional state. Currently, the major tools for determining metabolite concentrations are mass spectrometry coupled with chromatographic techniques and nuclear magnetic resonance, which are expensive, time consuming and destructive for the samples. Here, we report a time resolved approach to monitor metabolite dynamics in cell cultures, based on Surface Enhanced Raman Scattering (SERS). This method is label-free, easy to use and provides the opportunity to simultaneously study a broad range of molecules, without the need to process the biological samples. As proof of concept, NIH/3T3 cells were cultured in vitro, and the extracellular medium was collected at different time points to be analyzed with our engineered SERS substrates. By identifying individual peaks of the Raman spectra, we showed the simultaneous detection of several components of the conditioned medium, such as L-tyrosine, L-tryptophan, glycine, L-phenylalanine, L-histidine and fetal bovine serum proteins, as well as their intensity changes during time. Furthermore, analyzing the whole Raman data set with the Principal Component Analysis (PCA), we demonstrated that the Raman spectra collected at different days of culture and clustered by similarity, described a well-defined trajectory in the principal component plot. This approach was then utilized to determine indirectly the functional state of the macrophage cell line Raw 264.7, stimulated with the lipopolysaccharide (LPS) for 24 hours. The collected spectra at different time points, clustered by the PCA analysis, followed a well-defined trajectory, corresponding to the functional change of cells toward the activated pro-inflammatory state induced by the LPS. This study suggests that our engineered SERS surfaces can be used as a versatile tool both for the characterization of cell culture conditions and the functional state of cells over time.

Suggested Citation

  • Victoria Shalabaeva & Laura Lovato & Rosanna La Rocca & Gabriele C Messina & Michele Dipalo & Ermanno Miele & Michela Perrone & Francesco Gentile & Francesco De Angelis, 2017. "Time resolved and label free monitoring of extracellular metabolites by surface enhanced Raman spectroscopy," PLOS ONE, Public Library of Science, vol. 12(4), pages 1-16, April.
  • Handle: RePEc:plo:pone00:0175581
    DOI: 10.1371/journal.pone.0175581
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    References listed on IDEAS

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    1. Taro Ichimura & Liang-da Chiu & Katsumasa Fujita & Satoshi Kawata & Tomonobu M Watanabe & Toshio Yanagida & Hideaki Fujita, 2014. "Visualizing Cell State Transition Using Raman Spectroscopy," PLOS ONE, Public Library of Science, vol. 9(1), pages 1-8, January.
    2. Arun Sreekumar & Laila M. Poisson & Thekkelnaycke M. Rajendiran & Amjad P. Khan & Qi Cao & Jindan Yu & Bharathi Laxman & Rohit Mehra & Robert J. Lonigro & Yong Li & Mukesh K. Nyati & Aarif Ahsan & Sha, 2009. "Metabolomic profiles delineate potential role for sarcosine in prostate cancer progression," Nature, Nature, vol. 457(7231), pages 910-914, February.
    3. Giulia Rusciano & Paola Capriglione & Giuseppe Pesce & Salvatore Del Prete & Gilda Cennamo & David Di Cave & Luciano Cerulli & Antonio Sasso, 2013. "Raman Microspectroscopy Analysis in the Treatment of Acanthamoeba Keratitis," PLOS ONE, Public Library of Science, vol. 8(8), pages 1-8, August.
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