Analytical and Bioanalytical Chemistry

, Volume 411, Issue 3, pp 549–558 | Cite as

Quantitative mapping of specific proteins in biological tissues by laser ablation–ICP-MS using exogenous labels: aspects to be considered

  • María Cruz-Alonso
  • Ana Lores-Padín
  • Eva Valencia
  • Héctor González-IglesiasEmail author
  • Beatriz FernándezEmail author
  • Rosario Pereiro
Part of the following topical collections:
  1. Elemental and Molecular Imaging by LA-ICP-MS


Laser ablation (LA) coupled with inductively coupled plasma mass spectrometry (ICP-MS) is a versatile tool for direct trace elemental and isotopic analysis of solids. The development of new strategies for quantitative elemental mapping of biological tissues is one of the growing research areas in LA-ICP-MS. On the other hand, the latest advances are related to obtaining not only the elemental distribution of heteroatoms but also molecular information. In this vein, mapping of specific proteins in biological tissues can be done with LA-ICP-MS by use of metal-labelled immunoprobes. However, although LA-ICP-MS is, in principle, a quantitative technique, critical requirements should be met for absolute quantification of protein distribution. In this review, progress based on the use of metal-labelled antibodies for LA-ICP-MS mapping of specific proteins is reported. Critical requirements to obtain absolute quantitative mapping of specific proteins by LA-ICP-MS are highlighted. Additionally, illustrative examples of the advances made so far with LA-ICP-MS are provided.

Graphical abstract

In the proposed critical review, last advances based on the use of metal-labelled antibodies and critical requirements for LA-ICP-MS quantitative mapping of specific proteins are tackled.


Immunohistochemistry Antibody labelling Laser ablation Inductively coupled plasm mass spectrometry Protein quantification 



This work was supported through the project CTQ2016-79015-R by Agencia Estatal de Investigación (Spain) and FEDER. B.F. acknowledges her contract RYC-2014-14985 with the Spanish Ministry of Economy and Competitiveness through the Ramón y Cajal programme. The Instituto Oftalmológico Fernández-Vega and Fundación de Investigación Oftalmológica acknowledge support from Cátedra Rafael del Pino (Fundación Rafael del Pino), and Instituto de Desarrollo Económico del Principado de Asturias of Principado de Asturias and FEDER (project IDE/2016/000214).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.


  1. 1.
    Calderón-Celis F, Ruiz Encinar J, Sanz-Medel A. Standardization approaches in absolute quantitative proteomics with mass spectrometry. Mass Spectrom Rev. 2017:1–23.
  2. 2.
    Cid-Barrio L, Calderón-Celis F, Abásolo-Linares P, Fernández-Sánchez ML, Costa-Fernández JM, Ruiz Encinar J, et al. Advances in absolute protein quantification and quantitative protein mapping using ICP-MS. Trends Anal Chem. 2018;104:148–59. Scholar
  3. 3.
    Bishop DP, Cole N, Zhang T, Doble PA, Hare DJ. A guide to integrating immunohistochemistry and chemical imaging. Chem Soc Rev. 2018;47:3770–87. Scholar
  4. 4.
    Sancey L, Motto-Ros V, Busser B, Kotb S, Benoit JM, Piednoir A, et al. Laser spectrometry for multi-elemental imaging of biological tissues. Sci Rep. 2014;4:6065. Scholar
  5. 5.
    Rzagalinski I, Volmer DA. Quantification of low molecular weight compounds by MALDI imaging mass spectrometry – a tutorial review. Biochim Biophys Acta. 1865;2017:726–39. Scholar
  6. 6.
    Higashi T, Ogawa S. Isotope-coded ESI-enhancing derivatization reagents for differential analysis, quantification and profiling of metabolites in biological samples by LC/MS: a review. J Pharm Biomed Anal. 2016;130:181–93. Scholar
  7. 7.
    Sanz-Medel A, Montes-Bayón M, Bettmer J, Fernández-Sanchez ML, Ruiz EJ. ICP-MS for absolute quantification of proteins for heteroatom-tagged, targeted proteomics. Trends Anal Chem. 2012;40:52–63. Scholar
  8. 8.
    Jakubowski N, Messerschmidt J, Anorbe MG, Waentig L, Hayen H, Roos PH. Labelling of proteins by use of iodination and detection by ICP-MS. J Anal At Spectrom. 2008;23:1487–96. Scholar
  9. 9.
    Bomke S, Pfeifer T, Meermann B, Busche W, Karst U. Liquid chromatography with complementary electrospray and inductively coupled plasma mass spectrometric detection of ferrocene labelled peptides and proteins. Anal Bioanal Chem. 2010;397:3503–13. Scholar
  10. 10.
    Kutscher DJ, del Castillo Busto ME, Zinn N, Sanz-Medel A, Bettmer J. Protein labelling with mercury tags: fundamental studies on ovalbumin derivatised with p-hydroxymercuribenzoic acid (pHMB). J Anal At Spectrom. 2008;23:1359–1364.
  11. 11.
    de Bang TC, Husted S. Lanthanide elements as labels for multiplexed and targeted analysis of proteins, DNA and RNA using inductively-coupled plasma mass spectrometry. Trends Anal Chem. 2015;72:45–52.
  12. 12.
    El-Khatib AH, Esteban-Fernández D, Linscheid MW. Dual labeling of biomolecules using MeCAT and DOTA derivatives: application to quantitative proteomics. Anal Bioanal Chem. 2012;403:2255–67. Scholar
  13. 13.
    Zhang C, Wu F, Zhang Y, Wang X, Zhang X. A novel combination of immunoreaction and ICP-MS as a hyphenated technique for the determination of thyroid-stimulating hormone (TSH) in human serum. J Anal At Spectrom. 2001;16:1393–6. Scholar
  14. 14.
    Zhang C, Zhang Z, Yu B, Shi J, Zhang XR. Application of the biological conjugate between antibody and colloid Au nanoparticles as analyte to inductively coupled plasma mass spectrometry. Anal Chem. 2002;74:96–9. Scholar
  15. 15.
    Baranov VI, Quinn Z, Bandura DR, Tanner SD. A sensitive and quantitative element-tagged immunoassay with ICPMS detection. Anal Chem. 2002;74:1629–36. Scholar
  16. 16.
    Bradbury A, Plückthun A. Reproducibility: standardize antibodies used in research. Nature. 2015;518(7537):27–9. Scholar
  17. 17.
    Careri M, Elviri L, Mangia A. Element-tagged immunoassay with inductively coupled plasma mass spectrometry for multianalyte detection. Anal Bioanal Chem. 2009;393:57–61. Scholar
  18. 18.
    Liu Z, Li X, Xiao G, Chen B, He M, Hu B. Application of inductively coupled plasma mass spectrometry in the quantitative analysis of biomolecules with exogenous tags: a review. Trends Anal Chem. 2017;93:78–101. Scholar
  19. 19.
    Tanner SD, Baranov VI, Ornatsky OI, Bandura DR, George TC. An introduction to mass cytometry: fundamentals and applications. Cancer Immunol Immunother. 2013;62:955–65. Scholar
  20. 20.
    Mueller L, Traub H, Jakubowski N, Drescher D, Baranov VI, Kneipp J. Trends in single-cell analysis by use of ICP-MS. Anal Bioanal Chem. 2014;406:6963–77. Scholar
  21. 21.
    Schwarz G, Mueller L, Beck S, Linscheid MW. DOTA based metal labels for protein quantification: a review. J Anal At Spectrom. 2014;29:221–33. Scholar
  22. 22.
    Majonis D, Herrera I, Ornatsky O, Schulze M, Lou X, Soleimani M, et al. Synthesis of a functional metal-chelating polymer and steps toward quantitative mass cytometry bioassays. Anal Chem. 2010;82:8961–9. Scholar
  23. 23.
    Liu J-M, Yan X-P. Ultrasensitive, selective and simultaneous detection of cytochrome c and insulin based on immunoassay and aptamer-based bioassay in combination with Au/Ag nanoparticle tagging and ICP-MS detection. J Anal At Spectrom. 2011;26:1191–7. Scholar
  24. 24.
    Cruz-Alonso M, Fernandez B, Álvarez L, González-Iglesias H, Traub H, Jakubowski N, et al. Bioimaging of metallothioneins in ocular tissue sections by LA-ICP-MS using bioconjugated gold nanoclusters as specific tags. Microchim Acta. 2018;185:64. Scholar
  25. 25.
    Limbeck A, Galler P, Bonta M, Bauer G, Nischkauer W, Vanhaecke F. Recent advances in quantitative LA-ICP-MS analysis: challenges and solutions in the life sciences and environmental chemistry. Anal Bioanal Chem. 2015;407:6593–617. Scholar
  26. 26.
    Wang HAO, Grolimund D, Giesen C, Borca CN, Shaw-Stewart JRH, Bodenmiller B, et al. Fast chemical imaging at high spatial resolution by laser ablation inductively coupled plasma mass spectrometry. Anal Chem. 2013;85:10107–16. Scholar
  27. 27.
    Van Malderen SJM, Managh AJ, Sharp BL, Vanhaecke F. Recent developments in the design of rapid response cells for laser ablation-inductively coupled plasma-mass spectrometry and their impact on bioimaging applications. J Anal At Spectrom. 2016;31:423–439.
  28. 28.
    Chang Q, Ornatsky OI, Siddiqui I, Loboda A, Baranov VI, Hedley DW. Imaging mass cytometry. Cytometry Part A. 2017;91A:160–9. Scholar
  29. 29.
    Giesen C, Wang HAO, Schapiro D, Zivanovic N, Jacobs A, Hattendorf B, et al. Highly multiplexed imaging of tumor tissues with subcellular resolution by mass cytometry. Nat Methods. 2014;11:417–22. Scholar
  30. 30.
    Konz I, Fernández B, Fernández ML, Pereiro R, Sanz-Medel A. Laser ablation ICP-MS for quantitative biomedical applications. Anal Bioanal Chem. 2012;403:2113–25. Scholar
  31. 31.
    Pozebon D, Scheffler GL, Dressler VL. Recent applications of laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) for biological sample analysis: a follow-up review. J Anal At Spectrom. 2017;32:890–919. Scholar
  32. 32.
    Sussulini A, Becker JS, Becker JS. Laser ablation ICP-MS: application in biomedical research. Mass Spectrom Rev. 2017;36:47–57. Scholar
  33. 33.
    Stack EC, Wang C, Roman KA, Hoyt CC. Multiplexed immunohistochemistry, imaging, and quantitation: a review, with an assessment of tyramide signal amplification, multispectral imaging and multiplex analysis. Methods. 2014;70(1):46–58. Scholar
  34. 34.
    Matos LL, Trufelli DC, de Matos MG, da Silva Pinhal MA. Immunohistochemistry as an important tool in biomarkers detection and clinical practice. Biomark Insights. 2010;5:9–20.Google Scholar
  35. 35.
    Rodríguez-Menéndez S, Fernández B, García M, Álvarez L, Fernández ML, Sanz-Medel A, et al. Quantitative study of zinc and metallothioneins in the human retina and RPE cells by mass spectrometry-based methodologies. Talanta. 2018;178:222–30. Scholar
  36. 36.
    Taylor CR, Levenson RM. Quantification of immunohistochemistry-issues concerning methods, utility and semiquantitative assessment II. Histopathology. 2006;49:411–24. Scholar
  37. 37.
    Bonta M, Török S, Hegedus B, Döme B, Limbeck A. A comparison of sample preparation strategies for biological tissues and subsequent trace element analysis using LA-ICP-MS. Anal Bioanal Chem. 2017;409:1805–14. Scholar
  38. 38.
    Giesen C, Waentig L, Mairinger T, Drescher D, Kneipp J, Roos PH, et al. Iodine as an elemental marker for imaging of single cells and tissue sections by laser ablation inductively coupled plasma mass spectrometry. J Anal At Spectrom. 2011;26:2160–5. Scholar
  39. 39.
    Frick DA, Giesen C, Hemmerle T, Bodenmiller B, Günther D. An internal standardisation strategy for quantitative immunoassay tissue imaging using laser ablation inductively coupled plasma mass spectrometry. J Anal At Spectrom. 2015;30:254–9. Scholar
  40. 40.
    Hoesl S, Neumann B, Techritz S, Sauter G, Simon R, Schlüter H, et al. Internal standardization of LA-ICP-MS immune imaging via printing of universal metal spiked inks onto tissue sections. J Anal At Spectrom. 2016;31:801–8. Scholar
  41. 41.
    Mueller L, Herrmann AJ, Techritz S, Panne U, Jakubowski N. Quantitative characterization of single cells by use of immunocytochemistry combined with multiplex LA-ICP-MS. Anal Bioanal Chem. 2017;409:3667–76. Scholar
  42. 42.
    Boutureira O, Bernardes GJ. Advances in chemical protein modification. Chem Rev. 2015;115:2174–95. Scholar
  43. 43.
    de Gruyter JN, Malins LR, Baran PS. Residue-specific peptide modification: a chemist’s guide. Biochemistry. 2017;56:3863–73. Scholar
  44. 44.
    Giesen C, Mairinger T, Khoury L, Waentig L, Jakubowski N, Panne U. Multiplexed immunohistochemical detection of tumor markers in breast cancer tissue using laser ablation inductively coupled plasma mass spectrometry. Anal Chem. 2011;83:8177–83. Scholar
  45. 45.
    Ahrends R, Pieper S, Kühn A, Weisshoff H, Hamester M, Lindemann T, et al. A metal-coded affinity tag approach to quantitative proteomics. Mol Cell Proteomics. 2007;6:1907–16. Scholar
  46. 46.
    Schwarz G, Beck S, Weller MG, Linscheid MW. MeCAT—new iodoacetamide reagents for metal labelling of proteins and peptides. Anal Bioanal Chem. 2011;401:1203–9. Scholar
  47. 47.
    Waentig L, Roos PH, Jakubowski N. Labelling of antibodies and detection by laser ablation inductively coupled plasma mass spectrometry. PART III. Optimization of antibody labelling for application in a Western blot procedures. J Anal At Spectrom. 2009;24:924–33. Scholar
  48. 48.
    Waentig L, Jakubowski N, Roos PH. Multi-parametric analysis of cytochrome P450 expression in rat liver microsomes by LA-ICP-MS. J Anal At Spectrom. 2011;26:310–9. Scholar
  49. 49.
    Waentig L, Jakubowski N, Hardt S, Scheler C, Roos PH, Linscheid MW. Comparison of different chelates for lanthanide labeling of antibodies and application in a Western blot immunoassay combined with detection by laser. J Anal At Spectrom. 2012;27:1311–20. Scholar
  50. 50.
    de Bang TC, Pedas P, Schjoerring JK, Jensen PE, Husted S. Multiplexed quantification of plant thylakoid proteins on western blots using lanthanide-labeled antibodies and laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS). Anal Chem. 2013;85:5047–54. Scholar
  51. 51.
    Waentig L, Techritz S, Jakubowski N, Roos PH. A multi-parametric microarray for protein profiling: simultaneous analysis of 8 different cytochromes via differentially element tagged antibodies and laser ablation ICP-MS. Analyst. 2013;138:6309–15. Scholar
  52. 52.
    Mueller L, Mairinger T, Hermann G, Koellensperger G, Hann S. Characterization of metal-tagged antibodies used in ICP-MS-based immunoassays. Anal Bioanal Chem. 2014;406:163–9. Scholar
  53. 53.
    Kanje S, Herrmann AJ, Hober S, Mueller L. Next generation of labelling reagents for quantitative and multiplexing immunoassays by the use of LA-ICP-MS. Analyst. 2016;141:6374–80. Scholar
  54. 54.
    Lou X, Zhang G, Herrera I, Kinach R, Ornatsky O, Baranov V, et al. Polymer-based elemental tags for sensitive bioassays. Angew Chem Int Ed. 2007;46:6111–4. Scholar
  55. 55.
    Müller SD, Diaz-Bone RA, Felix J, Goedecke W. Detection of specific proteins by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) using gold cluster labelled antibodies. J Anal At Spectrom. 2005;20:907–11. Scholar
  56. 56.
    Seuma J, Bunch J, Cox A, McLeod C, Bell J, Murray C. Combination of immunohistochemistry and laser ablation ICP mass spectrometry for imaging of cancer biomarkers. Proteomics. 2008;8:3775–84. Scholar
  57. 57.
    Hare DJ, Lei P, Ayton S, Roberts BR, Grimm R, George JL, et al. An iron–dopamine index predicts risk of parkinsonian neurodegeneration in the substantia nigra pars compacta. Chem Sci. 2014;5:2160–9. Scholar
  58. 58.
    Paul B, Hare DJ, Bishop DP, Paton C, Nguyen VT, Cole N, et al. Visualising mouse neuroanatomy and function by metal distribution using laser ablation-inductively coupled plasma-mass spectrometry imaging. Chem Sci. 2015;6:5383–93. Scholar
  59. 59.
    Li J, Zhu J-J, Xu K. Fluorescent metal nanoclusters: from synthesis to applications. Trend Anal Chem. 2014;58:90–8. Scholar
  60. 60.
    Zhang L, Wang E. Metal nanoclusters: New fluorescent probes for sensors and bioimaging. Nano Today. 2014;9:132–57. Scholar
  61. 61.
    Alonso MC, Trapiella-Alfonso L, Fernández JMC, Pereiro R, Sanz-Medel A. Functionalized gold nanoclusters as fluorescent labels for immunoassays: application to human serum immunoglobulin E determination. Biosens Bioelectron. 2016;77:1055–61. Scholar
  62. 62.
    Díez I, Ras RHA. Fluorescent silver nanoclusters. Nanoscale. 2011;3:1963–70. Scholar
  63. 63.
    Fernández JF, Trapiella-Alfonso L, Costa-Fernández JM, Pereiro R, Sanz-Medel A. Aqueous synthesis of near-infrared highly fluorescent platinum nanoclusters. Nanotechnology. 2015;26:215601. Scholar
  64. 64.
    Buckle T, van der Wal S, van Malderen SJM, Müller L, Kuil J, van Unen V, Peters RJB, van Bemmel MEM, McDonnell LA, Velders AH, Koning F, Vanhaeke F, van Leeuwen FWB. Hybrid imaging labels: providing the link between mass spectrometry-based molecular pathology and theranostics. Theranostics. 2017;7:624–633.
  65. 65.
    Yang B, Zhang Y, Chen B, He M, Yin X, Wang H, et al. A multifunctional probe for ICP-MS determination and multimodal imaging of cancer cells. Biosens Bioelectron. 2017;96:77–83. Scholar
  66. 66.
    Ruhe L, Ickert S, Beck S, Linscheid MW. A new strategy for metal labeling of glycan structures in antibodies. Anal Bioanal Chem. 2018;410:21–5. Scholar
  67. 67.
    Beck A, Goetsch G, Dumontet C, Corvaïa N. Strategies and challenges for the next generation of antibody–drug conjugates. Nat Rev Drug Discov. 2017;16:315–37. Scholar
  68. 68.
    Marquez BV, Ikotun OF, Zheleznyak A, Wright B, Hari-Raj A, Pierce RA, et al. Evaluation of 89Zr-pertuzumab in breast cancer xenografts. Mol Pharm. 2014;11:3988–95. Scholar
  69. 69.
    Zhang S, Zhang S, Zhang X, Liu R. Analysis of proteins and DNAs using inductively coupled plasma mass spectrometry and elemental tagging. In: Encyclopedia of analytical chemistry: applications, theory and instrumentation. 2018:1–45.

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • María Cruz-Alonso
    • 1
  • Ana Lores-Padín
    • 1
  • Eva Valencia
    • 1
  • Héctor González-Iglesias
    • 2
    • 3
    Email author
  • Beatriz Fernández
    • 1
    • 3
    Email author
  • Rosario Pereiro
    • 1
    • 3
  1. 1.Department of Physical and Analytical ChemistryUniversity of OviedoOviedoSpain
  2. 2.Instituto Oftalmológico Fernández-VegaOviedoSpain
  3. 3.Instituto Universitario Fernández-Vega, Fundación de Investigación OftalmológicaUniversidad de OviedoOviedoSpain

Personalised recommendations