Analytical and Bioanalytical Chemistry

, Volume 390, Issue 6, pp 1585–1594 | Cite as

Quantitative micro-analysis of metal ions in subcellular compartments of cultured dopaminergic cells by combination of three ion beam techniques

  • A. Carmona
  • G. Devès
  • R. OrtegaEmail author
Original Paper


Quantification of the trace element content of subcellular compartments is a challenging task because of the lack of analytical quantitative techniques with adequate spatial resolution and sensitivity. Ion beam micro-analysis, using MeV protons or alpha particles, offers a unique combination of analytical methods that can be used with micrometric resolution for the determination of chemical element distributions. This work illustrates how the association of three ion beam analytical methods, PIXE (particle induced X-ray emission), BS (backscattering spectrometry), and STIM (scanning transmission ion spectrometry), allows quantitative determination of the trace element content of single cells. PIXE is used for trace element detection while BS enables beam-current normalization, and STIM local mass determination. These methods were applied to freeze-dried cells, following a specific cryogenic protocol for sample preparation which preserves biological structures and chemical distributions in the cells. We investigated how iron accumulates into dopaminergic cells cultured in vitro. We found that the iron content increases in dopaminergic cells exposed to an excess iron, with marked accumulation within distal ends, suggesting interaction between iron and dopamine within neurotransmitter vesicles. Increased iron content of dopaminergic neurons is suspected to promote neurodegeneration in Parkinson’s disease.


Ion beam analysis PIXE RBS Trace element Cell imaging 



The development of these experiments was supported by the European program of integrated action “Picasso”. The authors are grateful to Professor J. Lopez-Barneo, University of Sevilla, Spain, for providing the PC12 cell line. The authors are also grateful to the staff from CNA and CENBG, and especially Professor Ph. Moretto and P. Castel.


  1. 1.
    Yuste D (2005) Nat Methods 2:902–904CrossRefGoogle Scholar
  2. 2.
    Giepmans BN, Adams SR, Ellisman MH, Tsien RY (2006) Science 312:217–224CrossRefGoogle Scholar
  3. 3.
    Lobinski R, Moulin C, Ortega R (2006) Biochimie 88:1591–1604CrossRefGoogle Scholar
  4. 4.
    Cuajungco MP, Faget KY (2003) Brain Res Rev 41:44–56CrossRefGoogle Scholar
  5. 5.
    Bossy-Wetzel E, Schwarzenbacher R, Lipton SA (2004) Nat Med 10:S2–S9CrossRefGoogle Scholar
  6. 6.
    Gaeta A, Hider RC (2005) Br J Pharmacol 146:1041–1059CrossRefGoogle Scholar
  7. 7.
    Wolozin B, Golts N (2002) Neuroscientist 8:22–32CrossRefGoogle Scholar
  8. 8.
    Ke Y, Ming Qian Z (2003) Lancet Neurol 2:246–253CrossRefGoogle Scholar
  9. 9.
    Zecca L, Youdim MB, Riederer P, Connor JR, Crichton RR (2004) Nat Rev Neurosci 5:863–873CrossRefGoogle Scholar
  10. 10.
    Ortega R, Moretto Ph, Fajac A, Bénard J, Llabador Y, Simonoff M (1996) Cell Mol Biol 42:77–88Google Scholar
  11. 11.
    Ortega R, Bohic S, Tucoulou R, Somogy A, Devès G (2004) Anal Chem 76:309–314CrossRefGoogle Scholar
  12. 12.
    Johansson SAE, Johansson Th (1976) Nucl Instrum Methods 137:473–516CrossRefGoogle Scholar
  13. 13.
    Johansson SAE, Campbell JL, MalmQvist KG (1995) Particle-induced X-ray emission spectrometry (PIXE) Chemical Analysis Series, vol 133, WileyGoogle Scholar
  14. 14.
    Chu WK, Mayer JW, Nicolet MA (1978) Backscattering spectrometry. Academic Press, Orlando, FLGoogle Scholar
  15. 15.
    Moretto Ph, Razafindrabe L (1995) Nucl Instrum Methods Phys Res B 104:171–175CrossRefGoogle Scholar
  16. 16.
    Grime GW (1996) Nucl Instrum Methods Phys Res B 109/110:170–174CrossRefGoogle Scholar
  17. 17.
    Schofield RMS, Lefevre HW, Overley JC, Macdonald JD (1988) Nucl Instrum Methods Phys Res B 30:398–403CrossRefGoogle Scholar
  18. 18.
    Devès G, Ortega R (2002) Anal Bioanal Chem 374:390–394CrossRefGoogle Scholar
  19. 19.
    Devès G, Cohen-Bouhacina T, Ortega R (2004) Spectrochim Acta B 59:1733–1738CrossRefGoogle Scholar
  20. 20.
    Llabador Y, Moretto Ph (1998) Nuclear microprobes in the life sciences. World Scientific Publishing, SingaporeGoogle Scholar
  21. 21.
    Devès G, Michelet-Habchi C, Ortega R (2005) Nucl Instrum Methods Phys Res B 231:136–141CrossRefGoogle Scholar
  22. 22.
    Maxwell JA, Campbell JL, Teesdale WJ (1989) Nucl Instrum Methods Phys Res B 43:218–230CrossRefGoogle Scholar
  23. 23.
    Maxwell JA, Teesdale WJ, Campbell JL (1995) Nucl Instrum Methods Phys Res B 95:407–421CrossRefGoogle Scholar
  24. 24.
    Campbell JL, Hopman TL, Maxwell JA, Nejedly Z (2000) Nucl Instrum Methods Phys Res B 170:193–204CrossRefGoogle Scholar
  25. 25.
    Mayer M (1997) SIMNRA User’s Guide. Report IPP 9/113, Max-Planck-Institut für Plasmaphysik, Garching, GermanyGoogle Scholar
  26. 26.
    Greene LA, Tischler AS (1976) Proc Natl Acad Sci USA 73:2424–2428CrossRefGoogle Scholar
  27. 27.
    Tischler AS, Greene LA (1978) Lab Invest 39:77–89Google Scholar
  28. 28.
    Das KP, Freudenrich TM, Mundy WR (2004) Neurotoxicol Teratol 26:397–406CrossRefGoogle Scholar
  29. 29.
    Sulzer D, Bogulavsky J, Larsen KE, Behr G, Karatekin E, Kleinman MH, Turro N, Krantz D, Edwards RH, Greene LA, Zecca L (2000) Proc Natl Acad Sci 97:11869–11874CrossRefGoogle Scholar
  30. 30.
    Galvani P, Colleoni M, Origgi M, Santagostino A (1995) Toxicol In Vitro 9:365–368CrossRefGoogle Scholar
  31. 31.
    Watt F, Thong PSP, Tan AHM, Tang SM (1997) Nucl Instrum Methods Phys Res B 130:188–191CrossRefGoogle Scholar
  32. 32.
    Snyder WS, Cook MJ, Nasset ES, Karhausen LR, Parry Howells GP, Tipton IH (1975) Report of the task group on reference man, ICRP publication 23. Pergamon Press, OxfordGoogle Scholar
  33. 33.
    Nejedly Z, Campbell JL (2000) Nucl Instrum Methods Phys Res B 160:415–423CrossRefGoogle Scholar
  34. 34.
    Maenhaut W, Raemdonck H (1984) Nucl Instrum Methods Phys Res B 1:123–136CrossRefGoogle Scholar
  35. 35.
    Devès G, Ortega R (2001) Nucl Instrum Methods Phys B 181:460–464CrossRefGoogle Scholar
  36. 36.
    Maetz M, Przybylowicz WJ, Mesjasz-Przybylowicz J, Shübler A, Traxel K (1999) Nucl Instrum Methods Phys Res B 158:292–298CrossRefGoogle Scholar
  37. 37.
    Oakley AE, Collingwood JF, Dobson J, Love G, Perrott HR, Edwardson JA, Elstne, M, Morris CM (2007) Neurology 68:1820–1825CrossRefGoogle Scholar
  38. 38.
    Ortega R, Cloetens P, Devès G, Carmona A, Bohic S (2007) PLoS ONE 2(9):e925CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  1. 1.Groupe d’Imagerie Chimique Cellulaire et Spéciation, Chemin du solarium, Laboratoire de Chimie Nucléaire Analytique et BioenvironnementaleUniversités de Bordeaux 1 & 2GradignanFrance
  2. 2.Groupe d’Imagerie Chimique Cellulaire et Spéciation, Chemin du solariumCNRS, Laboratoire de Chimie Nucléaire Analytique et BioenvironnementaleGradignanFrance
  3. 3.Centro Nacional de AceleradoresUniversidad de SevillaSevillaSpain

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