Skip to main content
SpringerLink
Log in

Biomolecular Imaging with a C60-SIMS/MALDI Dual Ion Source Hybrid Mass Spectrometer: Instrumentation, Matrix Enhancement, and Single Cell Analysis

  • Research Article
  • Published:
Journal of The American Society for Mass Spectrometry

Abstract

We describe a hybrid MALDI/C60-SIMS Q-TOF mass spectrometer and corresponding sample preparation protocols to image intact biomolecules and their fragments in mammalian spinal cord, individual invertebrate neurons, and cultured neuronal networks. A lateral spatial resolution of 10 μm was demonstrated, with further improvement feasible to 1 μm, sufficient to resolve cell outgrowth and interconnections in neuronal networks. The high mass resolution (>13,000 FWHM) and tandem mass spectrometry capability of this hybrid instrument enabled the confident identification of cellular metabolites. Sublimation of a suitable matrix, 2,5-dihydroxybenzoic acid, significantly enhanced the ion signal intensity for intact glycerophospholipid ions from mammalian nervous tissue, facilitating the acquisition of high-quality ion images for low-abundance biomolecules. These results illustrate that the combination of C60-SIMS and MALDI mass spectrometry offers particular benefits for studies that require the imaging of intact biomolecules with high spatial and mass resolution, such as investigations of single cells, subcellular organelles, and communities of cells.

ᅟ

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6

Similar content being viewed by others

References

  1. Rubakhin, S.S., Sweedler, J.V. Mass Spectrometry Imaging. In: Rubakhin S.S., Sweedler J.V., (Eds.), pp. 21–49. Humana Press: Totowa (2010)

  2. Setou, M.: Imaging Mass Spectrometry: Protocols for Mass Microscopy, pp. 1–257. Springer, Tokyo (2010)

  3. Chughtai, K., Heeren, R.M.A.: Mass spectrometric imaging for biomedical tissue analysis. Chem. Rev. 110, 3237–3277 (2010)

    Article  CAS  Google Scholar 

  4. Schwamborn, K., Caprioli, R.M.: Innovation molecular imaging by mass spectrometry - looking beyond classical histology. Nat. Rev. Cancer 10, 639–646 (2010)

    Article  CAS  Google Scholar 

  5. Cornett, D.S., Reyzer, M.L., Chaurand, P., Caprioli, R.M.: MALDI imaging mass spectrometry: Molecular snapshots of biochemical systems. Nat. Methods 4, 828–833 (2007)

    Article  CAS  Google Scholar 

  6. Rubakhin, S.S., Greenough, W.T., Sweedler, J.V.: Spatial profiling with MALDI MS: distribution of neuropeptides within single neurons. Anal. Chem. 75, 5374–5380 (2003)

    Article  CAS  Google Scholar 

  7. Rubakhin, S.S., Garden, R.W., Fuller, R.R., Sweedler, J.V.: Measuring the peptides in individual organelles with mass spectrometry. Nat. Biotechnol. 18, 172–175 (2000)

    Article  CAS  Google Scholar 

  8. Rubakhin, S.S., Sweedler, J.V.: Quantitative measurements of cell−cell signaling peptides with single-cell MALDI MS. Anal. Chem. 80, 7128–7136 (2008)

    Article  CAS  Google Scholar 

  9. Greving, M.P., Patti, G.J., Siuzdak, G.: Nanostructure-initiator mass spectrometry metabolite analysis and imaging. Anal. Chem. 83, 2–7 (2011)

    Article  CAS  Google Scholar 

  10. Eberlin, L.S., Ferreira, C.R., Dill, A.L., Ifa, D.R., Cooks, R.G.: Desorption electrospray ionization mass spectrometry for lipid characterization and biological tissue imaging. Biochim. Biophys. Acta 1811, 946–960 (2011)

    Article  CAS  Google Scholar 

  11. Eberlin, L.S., Ifa, D.R., Wu, C., Cooks, R.G.: Three-dimensional vizualization of mouse brain by lipid analysis using ambient ionization mass spectrometry. Angew. Chem. Int. Ed. 49, 873–876 (2010)

    Article  CAS  Google Scholar 

  12. Nemes, P., Woods, A.S., Vertes, A.: Simultaneous imaging of small metabolites and lipids in rat brain tissues at atmospheric pressure by laser ablation electrospray ionization mass spectrometry. Anal. Chem. 82, 982–988 (2010)

    Article  CAS  Google Scholar 

  13. Nemes, P., Barton, A.A., Vertes, A.: Three-dimensional imaging of metabolites in tissues under ambient conditions by laser ablation electrospray ionization mass spectrometry. Anal. Chem. 81, 6668–6675 (2009)

    Article  CAS  Google Scholar 

  14. Shrestha, B., Patt, J.M., Vertes, A.: In situ cell-by-cell imaging and analysis of small cell populations by mass spectrometry. Anal. Chem. 83, 2947–2955 (2011)

    Article  CAS  Google Scholar 

  15. Laskin, J., Heath, B.S., Roach, P.J., Cazares, L., Semmes, O.J.: Tissue imaging using nanospray desorption electrospray ionization mass spectrometry. Anal. Chem. 84, 141–148 (2011)

    Article  Google Scholar 

  16. Watrous, J., Roach, P., Heath, B., Alexandrov, T., Laskin, J., Dorrestein, P.C.: Metabolic profiling directly from the petri dish using nanospray desorption electrospray ionization imaging mass spectrometry. Anal. Chem. 85, 10385–10391 (2013)

    Article  CAS  Google Scholar 

  17. Nemes, P., Vertes, A.: Ambient mass spectrometry for in vivo local analysis and in situ molecular tissue imaging. TraC Trends Anal. Chem 34, 22–34 (2012)

    Article  CAS  Google Scholar 

  18. Harris, G.A., Galhena, A.S., Fernandez, F.M.: Ambient sampling/ionization mass spectrometry: applications and current trends. Anal. Chem. 83, 4508–4538 (2011)

    Article  CAS  Google Scholar 

  19. Ifa, D.R., Wu, C.P., Ouyang, Z., Cooks, R.G.: Desorption electrospray ionization and other ambient ionization methods: current progress and preview. Analyst 135, 669–681 (2010)

    Article  CAS  Google Scholar 

  20. Boxer, S.G., Kraft, M.L., Weber, P.K.: Advances in imaging secondary ion mass spectrometry for biological samples. Annu. Rev. Biophys. 38, 53–74 (2009)

    Article  CAS  Google Scholar 

  21. Lanni, E.J., Rubakhin, S.S., Sweedler, J.V.: Mass spectrometry imaging and profiling of single cells. J. Proteome 75, 5036–5051 (2012)

    Article  CAS  Google Scholar 

  22. CAMECA (2010) NanoSIMS 50/50 l. SIMS microprobe for ultra fine feature analysis. Available at: http://www.cameca.fr/instruments-for-research/nanosims.aspx. Accessed 4/3/2014.

  23. Kollmer, F., Paul, W., Krehl, M., Niehuis, E.: Ultra high spatial resolution SIMS with cluster ions—approaching the physical limits. Surf. Interface Anal. 45, 312–314 (2012)

    Article  Google Scholar 

  24. Fletcher, J.S., Lockyer, N.P., Vaidyanathan, S., Vickerman, J.C.: TOF-SIMS 3D biomolecular imaging of xenopus laevis oocytes using buckminsterfullerene (C60) primary ions. Anal. Chem. 29, 2199–2206 (2007)

    Article  Google Scholar 

  25. Fletcher, J.S., Vickerman, J.C., Winograd, N.: Label free biochemical 2D and 3D imaging using secondary ion mass spectrometry. Curr. Opin. Chem. Biol. 15, 733–740 (2011)

    Article  CAS  Google Scholar 

  26. Jones, E., Lockyer, N., Kordys, J., Vickerman, J.: Suppression and enhancement of secondary ion formation due to the chemical environment in static-secondary ion mass spectrometry. J. Am. Soc. Mass Spectrom. 18, 1559–1567 (2007)

    Article  CAS  Google Scholar 

  27. Kollmer, F.: Cluster primary ion bombardment of organic materials. Appl. Surf. Sci. 231/232, 153–158 (2004)

    Article  Google Scholar 

  28. Boussofiane-Baudin, K., Bolbach, G., Brunelle, A., Della-Negra, S., Håkansson, P., Le Beyec, Y.: Secondary ion emission under cluster impact at low energies (5–60 kev); influence of the number of atoms in the projectile. Nucl. Instrum. Meth. Phys. Res. B 88, 160–163 (1994)

    Article  CAS  Google Scholar 

  29. Matsuo, J., Okubo, C., Seki, T., Aoki, T., Toyoda, N., Yamada, I.: A new secondary ion mass spectrometry (SIMS) system with high-intensity cluster ion source. Nucl. Instrum. Meth. Phys. Res. B 219–220, 463–467 (2004)

    Article  Google Scholar 

  30. Carado, A., Passarelli, M.K., Kozole, J., Wingate, J.E., Winograd, N., Loboda, A.V.: C60 secondary ion mass spectrometry with a hybrid-quadrupole orthogonal time-of-flight mass spectrometer. Anal. Chem. 80, 7921–7929 (2008)

    Article  CAS  Google Scholar 

  31. Smith, D.F., Robinson, E.W., Tolmachev, A.V., Heeren, R.M.A., Paša-Tolić, L.: C60 secondary ion Fourier transform ion cyclotron resonance mass spectrometry. Anal. Chem. 83, 9552–9557 (2011)

    Article  CAS  Google Scholar 

  32. Fletcher, J.S., Rabbani, S., Henderson, A., Blenkinsopp, P., Thompson, S.P., Lockyer, N.P., Vickerman, J.C.: A new dynamic in mass spectral imaging of single biological cells. Anal. Chem. 80, 9058–9064 (2008)

    Article  CAS  Google Scholar 

  33. Passarelli, M.K., Ewing, A.G., Winograd, N.: C(60)-SIMS studies of glycerophospholipid in a lipid maps model system: kdo(2)-lipid a stimulated raw 264.7 cells. Surf. Interface Anal. 45, 298–301 (2013)

    Article  CAS  Google Scholar 

  34. Passarelli, M.K., Ewing, A.G., Winograd, N.: Single-cell lipidomics: characterizing and imaging lipids on the surface of individual Aplysia californica neurons with cluster secondary ion mass spectrometry. Anal. Chem. 85, 2231–2238 (2013)

    Article  CAS  Google Scholar 

  35. Willingham, D., Brenes, D.A., Winograd, N., Wucher, A.: Investigating the fundamentals of molecular depth profiling using strong-field photoionization of sputtered neutrals. Surf. Interface Anal. 43, 45–48 (2011)

    Article  CAS  Google Scholar 

  36. Winograd, N.: Ion beams and laser postionization for molecule-specific imaging. Anal. Chem. 65, 622A–629A (1993)

    Article  CAS  Google Scholar 

  37. Wucher, A., Tian, H., Winograd, N.: A mixed cluster ion beam to enhance the ionization efficiency in molecular secondary ion mass spectrometry. Rapid Commun. Mass Spectrom. 28, 396–400 (2014)

    Article  CAS  Google Scholar 

  38. Liao, H.-Y., Lin, K.-Y., Kao, W.-L., Chang, H.-Y., Huang, C.-C., Shyue, J.-J.: Enhancing the sensitivity of molecular secondary ion mass spectrometry with C60+-o2+ cosputtering. Anal. Chem. 85, 3781–3788 (2013)

    Article  CAS  Google Scholar 

  39. Grade, H., Cooks, R.G.: Secondary ion mass spectrometry. Cationization of organic molecules with metals. J. Am. Chem. Soc. 100, 5615–5621 (1978)

    Article  CAS  Google Scholar 

  40. Altelaar, A.F.M., Klinkert, I., Jalink, K., de Lange, R.P.J., Adan, R.A.H., Heeren, R.M.A., Piersma, S.R.: Gold-enhanced biomolecular surface imaging of cells and tissue by SIMS and MALDI mass spectrometry. Anal. Chem. 78, 734–742 (2005)

    Article  Google Scholar 

  41. Wu, K.J., Odom, R.W.: Matrix-enhanced secondary ion mass spectrometry: a method for molecular analysis of solid surfaces. Anal. Chem. 68, 873–882 (1996)

    Article  CAS  Google Scholar 

  42. Nicola, A., Muddiman, D., Hercules, D.: Enhancement of ion intensity in time-of-flight secondary-ionization mass spectrometry. J. Am. Soc. Mass Spectrom. 7, 467–472 (1996)

    Article  CAS  Google Scholar 

  43. Altelaar, A.F.M., Luxembourg, S.L., McDonnell, L.A., Piersma, S.R., Heeren, R.M.A.: Imaging mass spectrometry at cellular length scales. Nat. Protoc. 2, 1185–1196 (2007)

    Article  CAS  Google Scholar 

  44. Altelaar, A.F.M., Taban, I.M., McDonnell, L.A., Verhaert, P.D.E.M., de Lange, R.P.J., Adan, R.A.H., Mooi, W.J., Heeren, R.M.A., Piersma, S.R.: High-resolution MALDI imaging mass spectrometry allows localization of peptide distributions at cellular length scales in pituitary tissue sections. Int. J. Mass Spectrom. 260, 203–211 (2007)

    Article  CAS  Google Scholar 

  45. Wehbe, N., Heile, A., Arlinghaus, H.F., Bertrand, P., Delcorte, A.: Effects of metal nanoparticles on the secondary ion yields of a model alkane molecule upon atomic and polyatomic projectiles in secondary ion mass spectrometry. Anal. Chem. 80, 6235–6244 (2008)

    Article  CAS  Google Scholar 

  46. Delcorte, A., Yunus, S., Wehbe, N., Nieuwjaer, N., Poleunis, C., Felten, A., Houssiau, L., Pireaux, J.J., Bertrand, P.: Metal-assisted secondary ion mass spectrometry using atomic (ga+, in+) and fullerene projectiles. Anal. Chem. 79, 3673–3689 (2007)

    Article  CAS  Google Scholar 

  47. Locklear, J.E., Guillermier, C., Verkhoturov, S.V., Schweikert, E.A.: Matrix-enhanced cluster-SIMS. Appl. Surf. Sci. 252, 6624–6627 (2006)

    Article  CAS  Google Scholar 

  48. McDonnell, L.A., Heeren, R.M.A., de Lange, R.P.J., Fletcher, I.W.: Higher sensitivity secondary ion mass spectrometry of biological molecules for high resolution, chemically specific imaging. J. Am. Soc. Mass Spectrom. 17, 1195–1202 (2006)

    Article  CAS  Google Scholar 

  49. Monroe, E.B., Jurchen, J.C., Lee, J., Rubakhin, S.S., Sweedler, J.V.: Vitamin E imaging and localization in the neuronal membrane. J. Am. Chem. Soc. 127, 12152–12153 (2005)

    Article  CAS  Google Scholar 

  50. Hankin, J., Barkley, R., Murphy, R.: Sublimation as a method of matrix application for mass spectrometric imaging. J. Am. Soc. Mass Spectrom. 18, 1646–1652 (2007)

    Article  CAS  Google Scholar 

  51. Ryan, K.E., Smiley, E.J., Winograd, N., Garrison, B.J.: Angle of incidence effects in a molecular solid. Appl. Surf. Sci. 255, 844–846 (2008)

    Article  CAS  Google Scholar 

  52. Taormina, C.R., Nicolette, E., Pedder, R.E., Novak, T.T.: An ion guide study: quadrupoles, rectilinear quadrupoles, hexapoles, and octopoles, 2009. Available at: http://www.ardaratech.com/document-library/technical-notes/11-tn-3007-practical-quadrupole-theory-ion-guides. Accessed 8 Aug 2014

  53. Loboda, A.V., Krutchinsky, A.N., Bromirski, M., Ens, W., Standing, K.G.: A tandem quadrupole/time-of-flight mass spectrometer with a matrix-assisted laser desorption/ionization source: design and performance. Rapid Commun. Mass Spectrom. 14, 1047–1057 (2000)

    Article  CAS  Google Scholar 

  54. Brison, J., Robinson, M.A., Benoit, D.S., Muramoto, S., Stayton, P.S., Castner, D.G.: TOF-SIMS 3D imaging of native and non-native species within hela cells. Anal. Chem. 85, 10869–10877 (2013)

    Article  CAS  Google Scholar 

  55. Passarelli, M.K., Winograd, N.: Characterizing in situ glycerophospholipids with SIMS and MALDI methodologies. Surf. Interface Anal. 43, 269–271 (2011)

    Article  CAS  Google Scholar 

  56. Tucker, K.R., Li, Z., Rubakhin, S.S., Sweedler, J.V.: Secondary ion mass spectrometry imaging of molecular distributions in cultured neurons and their processes: comparative analysis of sample preparation. J. Am. Soc. Mass Spectrom. 23, 1931–1938 (2012)

    Article  CAS  Google Scholar 

  57. Miyamoto, M.D.: Binomial analysis of quantal transmitter release at glycerol treated frog neuromuscular junctions. J. Physiol. 250, 121–142 (1975)

    CAS  Google Scholar 

  58. Galle, P.: [Sur une nouvelle methode d'analyse cellulaire utilisant le phenomene d'emission ionique secondaire.]. Ann. Phys. Biol. Med. 42, 83–94 (1970)

    Google Scholar 

  59. Ostrowski, S.G., Van Bell, C.T., Winograd, N., Ewing, A.G.: Mass spectrometric imaging of highly curved membranes during Tetrahymena mating. Science 305, 71–73 (2004)

    Article  CAS  Google Scholar 

  60. Rabbani, S., Fletcher, J.S., Lockyer, N.P., Vickerman, J.C.: Exploring subcellular imaging on the buncher-ToF j105 3D chemical imager. Surf. Interface Anal. 43, 380–384 (2011)

    Article  CAS  Google Scholar 

  61. Tokareva, E.N., Fardim, P., Pranovich, A.V., Fagerholm, H.P., Daniel, G., Holmbom, B.: Imaging of wood tissue by ToF-SIMS: critical evaluation and development of sample preparation techniques. Appl. Surf. Sci. 253, 7569–7577 (2007)

    Article  CAS  Google Scholar 

  62. Robinson, M.A., Castner, D.G.: Characterization of sample preparation methods of nih/3t3 fibroblasts for ToF-SIMS analysis. Biointerphases 8, 15 (2013)

    Article  Google Scholar 

  63. Smith, C.A., O'Maille, G., Want, E.J., Qin, C., Trauger, S.A., Brandon, T.R., Custodio, D.E., Abagyan, R., Siuzdak, G.: Metlin: a metabolite mass spectral database. Ther. Drug Monit. 27, 747–751 (2005)

    Article  CAS  Google Scholar 

  64. Monroe, E.B., Annangudi, S.P., Hatcher, N.G., Gutstein, H.B., Rubakhin, S.S., Sweedler, J.V.: SIMS and MALDI MS imaging of the spinal cord. Proteomics 8, 3746–3754 (2008)

    Article  CAS  Google Scholar 

  65. Zingg, J.-M.: Modulation of signal transduction by vitamin E. Mol. Aspects Med. 28, 481–506 (2007)

    Article  CAS  Google Scholar 

  66. Fletcher, J., Kotze, H., Armitage, E., Lockyer, N., Vickerman, J.: Evaluating the challenges associated with time-of-fight secondary ion mass spectrometry for metabolomics using pure and mixed metabolites. Metabolomics 9, 535–544 (2013)

    Article  CAS  Google Scholar 

  67. Glanzman, D.L., Kandel, E.R., Schacher, S.: Identified target motor neuron regulates neurite outgrowth and synapse formation of Aplysia sensory neurons in vitro. Neuron 3, 441–450 (1989)

    Article  CAS  Google Scholar 

  68. Girod, M., Shi, Y., Cheng, J.-X., Cooks, R.G.: Desorption electrospray ionization imaging mass spectrometry of lipids in rat spinal cord. J. Am. Soc. Mass Spectrom. 21, 1177–1189 (2010)

    Article  CAS  Google Scholar 

  69. Yang, J., Caprioli, R.M.: Matrix sublimation/recrystallization for imaging proteins by mass spectrometry at high spatial resolution. Anal. Chem. 83, 5728–5734 (2011)

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The authors are very grateful for instrumental design and troubleshooting advice provided by Alexandre Loboda at AB SCIEX (presently at Fludigm Corporation), Randy Pedder at Ardara Technologies, and the Winograd research group at Penn State University, particularly with Professor Nicholas Winograd, Melissa Passarelli, Anita Durairaj, and Lauren Jackson. Design assistance by Zhen Li and Kevin Tucker, precision machining by Michael A. Harland and Roger Smith at the Machine Shop of the School of Chemical Sciences at the University of Illinois at Urbana-Champaign, and cell dissection and culture assistance by Xiying Wang and Callie (Croushore) Kindt are also acknowledged. This work was supported by the Department of Energy Office of Biological and Environmental Research through grant DE SC0006642, and The National Resource for Aplysia funded by PHS grant P40 OD010952. The authors declare no competing financial interests.

Author information

Authors and Affiliations

  1. Department of Chemistry and Beckman Institute for Advanced Science and Technology, University of Illinois at Urbana-Champaign, Urbana, IL, 61801, USA

    Eric J. Lanni, Sage J. B. Dunham, Peter Nemes, Stanislav S. Rubakhin & Jonathan V. Sweedler

  2. Department of Chemistry, The George Washington University, Washington, DC, 20052, USA

    Peter Nemes

Authors
  1. Eric J. Lanni
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sage J. B. Dunham
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Peter Nemes
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Stanislav S. Rubakhin
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Jonathan V. Sweedler
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jonathan V. Sweedler.

Electronic Supplementary Material

Below is the link to the electronic supplementary material.

ESM 1

(PDF 532 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Lanni, E.J., Dunham, S.J.B., Nemes, P. et al. Biomolecular Imaging with a C60-SIMS/MALDI Dual Ion Source Hybrid Mass Spectrometer: Instrumentation, Matrix Enhancement, and Single Cell Analysis. J. Am. Soc. Mass Spectrom. 25, 1897–1907 (2014). https://doi.org/10.1007/s13361-014-0978-9

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13361-014-0978-9

Key words

Search

Navigation