Skip to main content
SpringerLink
Log in

Combining Laser Capture Microdissection and Proteomics Techniques

  • Protocol
Clinical Proteomics

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 428))

Summary

Laser microdissection is an effective technique to harvest pure cell populations from complex tissue sections. In addition to using the microdissected cells in several DNA and RNA studies, it has been shown that the small number of cells obtained by this technique can also be used for proteomics analysis. Combining laser capture microdissection and different types of mass spectrometers opened ways to find and identify proteins that are specific for various cell types, tissues, and their morbid alterations. Although the combination of microdissection followed by the currently available techniques of proteomics has not yet reached the stage of genome wide representation of all proteins present in a tissue, it is a feasible way to find significant differentially expressed proteins in target tissues. Recent developments in mass spectrometric detection followed by proper statistics and bioinformatics enable to analyze the proteome of not more than 100–200 cells. Obviously, validation of result is essential. The present review describes and discusses the various methods developed to target cell populations of interest by laser microdissection, followed by analysis of their proteome.

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

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 159.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Abbreviations

LCM:

Laser Capture Microdissection

LMM:

Laser Microbeam Microdissection

LPC:

Laser Pressure Catapulting

2D PAGE:

Two-dimensional Polyacrylamide Gel Electrophoresis

2D DIGE:

Differential In-gel Electrophoresis

SDS:

Sodium Dodecyl Sulphate

MALDI-TOF/MS:

Matrix-assisted Laser Desorption/Ionization Time-of-flight Mass Spectrometry

MALDI-FTMS:

Matrix-assisted Laser Desorption/Ionization Fourier Transformer Mass Spectrometry

LC-ESI-MS/MS:

Liquid Chromatography-Electrospray Ionization Tandem Mass Spectrometry

HPLC:

High Performance Liquid Chromatography

SELDI-TOF:

Surface-enhanced Laser Desorption/Ionization Time-of-flight

ICAT:

Isotope-coded Affinity Tag

References

  1. Zhang, L., Zhou, W., Velculescu, V. E., Kern, S. E., Hruban, R. H., Hamilton, S. R., Vogelstein, B. and Kinzler, K. W. (1997) Gene expression profiles in normal and cancer cells. Science 276, 1268–1272.

    Article  PubMed  CAS  Google Scholar 

  2. Curran, S., McKay, J. A., McLeod, H. L. and Murray, G. I. (2000) Laser capture microscopy. Mol Pathol 53, 64–68.

    Article  PubMed  CAS  Google Scholar 

  3. Going, J. J. and Lamb, R. F. (1996) Practical histological microdissection for PCR analysis. J Pathol 179, 121–124.

    Article  PubMed  CAS  Google Scholar 

  4. Zhuang, Z., Bertheau, P., Emmert-Buck, M. R., Liotta, L. A., Gnarra, J., Linehan, W. M. and Lubensky, I. A. (1995) A microdissection technique for archival DNA analysis of specific cell populations in lesions <1 mm in size. Am J Pathol 146, 620–625.

    PubMed  CAS  Google Scholar 

  5. Shibata, D., Hawes, D., Li, Z. H., Hernandez, A. M., Spruck, C. H. and Nichols, P. W. (1992) Specific genetic analysis of microscopic tissue after selective ultraviolet radiation fractionation and the polymerase chain reaction. Am J Pathol 141, 539–543.

    PubMed  CAS  Google Scholar 

  6. Emmert-Buck, M. R., Bonner, R. F., Smith, P. D., Chuaqui, R. F., Zhuang, Z., Goldstein, S. R., Weiss, R. A. and Liotta, L. A. (1996) Laser capture microdissection. Science 274, 998–1001.

    Article  PubMed  CAS  Google Scholar 

  7. Wang, Y., Rudnick, P. A., Evans, E. L., Li, J., Zhuang, Z., Devoe, D. L., Lee, C. S. and Balgley, B. M. (2005) Proteome analysis of microdissected tumor tissue using a capillary isoelectric focusing-based multidimensional separation platform coupled with ESI-tandem MS. Anal Chem 77, 6549–6556.

    Article  PubMed  CAS  Google Scholar 

  8. Suarez-Quian, C. A., Goldstein, S. R., Pohida, T., Smith, P. D., Peterson, J. I., Wellner, E., Ghany, M. and Bonner, R. F. (1999) Laser capture microdissection of single cells from complex tissues. Biotechniques 26, 328–335.

    PubMed  CAS  Google Scholar 

  9. Espina, V., Milia, J., Wu, G., Cowherd, S. and Liotta, L. A. (2006) Laser capture microdissection. Methods Mol Biol 319, 213–229.

    Article  PubMed  CAS  Google Scholar 

  10. Schutze, K., Posl, H. and Lahr, G. (1998) Laser micromanipulation systems as universal tools in cellular and molecular biology and in medicine. Cell Mol Biol (Noisy-le-grand) 44, 735–746.

    CAS  Google Scholar 

  11. Gillespie, J. W., Gannot, G., Tangrea, M. A., Ahram, M., Best, C. J., Bichsel, V. E., Petricoin, E. F., Emmert-Buck, M. R. and Chuaqui, R. F. (2004) Molecular profiling of cancer. Toxicol Pathol 32(Suppl. 1), 67–71.

    Article  PubMed  CAS  Google Scholar 

  12. Niyaz, Y., Stich, M., Sagmuller, B., Burgemeister, R., Friedemann, G., Sauer, U., Gangnus, R. and Schutze, K. (2005) Noncontact laser microdissection and pressure catapulting: sample preparation for genomic, transcriptomic, and proteomic analysis. Methods Mol Med 114, 1–24.

    PubMed  CAS  Google Scholar 

  13. Ball, H. J. and Hunt, N. H. (2004) Needle in a haystack: microdissecting the proteome of a tissue. Amino Acids 27, 1–7.

    Article  PubMed  CAS  Google Scholar 

  14. Emmert-Buck, M. R., Gillespie, J. W., Paweletz, C. P., Ornstein, D. K., Basrur, V., Appella, E., Wang, Q. H., Huang, J., Hu, N., Taylor, P. and Petricoin, E. F. 3rd (2000) An approach to proteomic analysis of human tumors. Mol Carcinog 27, 158–165.

    Article  PubMed  CAS  Google Scholar 

  15. Lawrie, L. C., Curran, S., McLeod, H. L., Fothergill, J. E. and Murray, G. I. (2001) Application of laser capture microdissection and proteomics in colon cancer. Mol Pathol 54, 253–258.

    Article  PubMed  CAS  Google Scholar 

  16. Jones, M. B., Krutzsch, H., Shu, H., Zhao, Y., Liotta, L. A., Kohn, E. C. and Petricoin, E. F. 3rd (2002) Proteomic analysis and identification of new biomarkers and therapeutic targets for invasive ovarian cancer. Proteomics 2, 76–84.

    Article  PubMed  CAS  Google Scholar 

  17. Wulfkuhle, J. D., Sgroi, D. C., Krutzsch, H., McLean, K., McGarvey, K., Knowlton, M., Chen, S., Shu, H., Sahin, A., Kurek, R., Wallwiener, D., Merino, M. J., Petricoin, E. F. 3rd, Zhao, Y. and Steeg, P. S. (2002) Proteomics of human breast ductal carcinoma in situ. Cancer Res 62, 6740–6749.

    PubMed  CAS  Google Scholar 

  18. Shekouh, A. R., Thompson, C. C., Prime, W., Campbell, F., Hamlett, J., Herrington, C. S., Lemoine, N. R., Crnogorac-Jurcevic, T., Buechler, M. W., Friess, H., Neoptolemos, J. P., Pennington, S. R. and Costello, E. (2003) Application of laser capture microdissection combined with two-dimensional electrophoresis for the discovery of differentially regulated proteins in pancreatic ductal adenocarcinoma. Proteomics 3, 1988–2001.

    Article  PubMed  CAS  Google Scholar 

  19. Zhang, D. H., Tai, L. K., Wong, L. L., Sethi, S. K. and Koay, E. S. (2005) Proteomics of breast cancer: enhanced expression of cytokeratin19 in human epidermal growth factor receptor type 2 positive breast tumors. Proteomics 5, 1797–1805.

    Article  PubMed  CAS  Google Scholar 

  20. Ai, J., Tan, Y., Ying, W., Hong, Y., Liu, S., Wu, M., Qian, X. and Wang, H. (2006) Proteome analysis of hepatocellular carcinoma by laser capture microdissection. Proteomics 6, 538–546.

    Article  PubMed  CAS  Google Scholar 

  21. Zhou, G., Li, H., DeCamp, D., Chen, S., Shu, H., Gong, Y., Flaig, M., Gillespie, J. W., Hu, N., Taylor, P. R., Emmert-Buck, M. R., Liotta, L. A., Petricoin, E. F. 3rd and Zhao, Y. (2002) 2D differential in-gel electrophoresis for the identification of esophageal scans cell cancer-specific protein markers. Mol Cell Proteomics 1, 117–124.

    Article  PubMed  CAS  Google Scholar 

  22. Lee, J. R., Baxter, T. M., Yamaguchi, H., Wang, T. C., Goldenring, J. R. and Anderson, M. G. (2003) Differential protein analysis of spasomolytic polypeptide expressing metaplasia using laser capture microdissection and two-dimensional difference gel electrophoresis. Appl Immunohistochem Mol Morphol 11, 188–193.

    Article  PubMed  CAS  Google Scholar 

  23. Hudelist, G., Singer, C. F., Pischinger, K. I., Kaserer, K., Manavi, M., Kubista, E. and Czerwenka, K. F. (2006) Proteomic analysis in human breast cancer: identification of a characteristic protein expression profile of malignant breast epithelium. Proteomics 6, 1989–2002.

    Article  PubMed  CAS  Google Scholar 

  24. Shaw, J., Rowlinson, R., Nickson, J., Stone, T., Sweet, A., Williams, K. and Tonge, R. (2003) Evaluation of saturation labelling two-dimensional difference gel electrophoresis fluorescent dyes. Proteomics 3, 1181–1195.

    Article  PubMed  CAS  Google Scholar 

  25. Greengauz-Roberts, O., Stoppler, H., Nomura, S., Yamaguchi, H., Goldenring, J. R., Podolsky, R. H., Lee, J. R. and Dynan, W. S. (2005) Saturation labeling with cysteine-reactive cyanine fluorescent dyes provides increased sensitivity for protein expression profiling of laser-microdissected clinical specimens. Proteomics 5, 1746–1757.

    Article  PubMed  CAS  Google Scholar 

  26. Sitek, B., Potthoff, S., Schulenborg, T., Stegbauer, J., Vinke, T., Rump, L. C., Meyer, H. E., Vonend, O. and Stuhler, K. (2006) Novel approaches to analyse glomerular proteins from smallest scale mu rine and human samples using DIGE saturation labelling. Proteomics 6, 4337–4345.

    Article  PubMed  CAS  Google Scholar 

  27. Li, C., Hong, Y., Tan, Y. X., Zhou, H., Ai, J. H., Li, S. J., Zhang, L., Xia, Q. C., Wu, J. R., Wang, H. Y. and Zeng, R. (2004) Accurate qualitative and quantitative proteomic analysis of clinical hepatocellular carcinoma using laser capture microdissection coupled with isotope-coded affinity tag and two-dimensional liquid chromatography mass spectrometry. Mol Cell Proteomics 3, 399–409.

    Article  PubMed  CAS  Google Scholar 

  28. Gygi, S. P., Rist, B., Gerber, S. A., Turecek, F., Gelb, M. H. and Aebersold, R. (1999) Quantitative analysis of complex protein mixtures using isotope-coded affinity tags. Nat Biotechnol 17, 994–999.

    Article  PubMed  CAS  Google Scholar 

  29. Zang, L., Palmer Toy, D., Hancock, W. S., Sgroi, D. C. and Karger, B. L. (2004) Proteomic analysis of ductal carcinoma of the breast using laser capture microdissection, LC-MS, and 16O/18O isotopic labeling. J Proteome Res 3, 604–612.

    Article  PubMed  CAS  Google Scholar 

  30. Poznanovic, S., Wozny, W., Schwall, G. P., Sastri, C., Hunzinger, C., Stegmann, W., Schrattenholz, A., Buchner, A., Gangnus, R., Burgemeister, R. and Cahill, M. A. (2005) Differential radioactive proteomic analysis of microdissected renal cell carcinoma tissue by 54 cm isoelectric focusing in serial immobilized pH gradient gels. J Proteome Res 4, 2117–2125.

    Article  PubMed  CAS  Google Scholar 

  31. Neubauer, H., Clare, S. E., Kurek, R., Fehm, T., Wallwiener, D., Sotlar, K., Nordheim, A., Wozny, W., Schwall, G. P., Poznanovic, S., Sastri, C., Hunzinger, C., Stegmann, W., Schrattenholz, A. and Cahill, M. A. (2006) Breast cancer proteomics by laser capture microdissection, sample pooling, 54-cm IPG IEF, and differential iodine radioisotope detection. Electrophoresis 27, 1840–1852.

    Article  PubMed  CAS  Google Scholar 

  32. Preisler, J., Hu, P., Rejtar, T., Moskovets, E. and Karger, B. L. (2002) Capillary array electrophoresis-MALDI mass spectrometry using a vacuum deposition interface. Anal Chem 74, 17–25.

    Article  PubMed  CAS  Google Scholar 

  33. Bergstrom, S. K., Samskog, J. and Markides, K. E. (2003) Development of a poly(dimethylsiloxane) interface for on-line capillary column liquid chromatography-capillary electrophoresis coupled to sheathless electrospray ionization time-of-flight mass spectrometry. Anal Chem 75, 5461–5467.

    Article  PubMed  Google Scholar 

  34. Wu, S. L., Hancock, W. S., Goodrich, G. G. and Kunitake, S. T. (2003) An approach to the proteomic analysis of a breast cancer cell line (SKBR-3). Proteomics 3, 1037–1046.

    Article  PubMed  CAS  Google Scholar 

  35. Gozal, Y. M., Cheng, D., Duong, D. M., Lah, J. J., Levey, A. I. and Peng, J. (2006) Merger of laser capture microdissection and mass spectrometry: a window into the amyloid plaque proteome. Methods Enzymol 412, 77–93.

    Article  PubMed  CAS  Google Scholar 

  36. Guo, J., Colgan, T. J., DeSouza, L. V., Rodrigues, M. J., Romaschin, A. D. and Siu, K. W. (2005) Direct analysis of laser capture microdissected endometrial carcinoma and epithelium by matrix-assisted laser desorption/ionization mass spectrometry. Rapid Commun Mass Spectrom 19, 2762–2766.

    Article  PubMed  CAS  Google Scholar 

  37. de Groot, C. J., Steegers-Theunissen, R. P., Guzel, C., Steegers, E. A. and Luider, T. M. (2005) Peptide patterns of laser dissected human trophoblasts analyzed by matrix-assisted laser desorption/ionisation-time of flight mass spectrometry. Proteomics 5, 597–607.

    Article  PubMed  Google Scholar 

  38. Umar, A., Dalebout, J. C., Timmermans, A. M., Foekens, J. A. and Luider, T. M. (2005) Method optimisation for peptide profiling of microdissected breast carcinoma tissue by matrix-assisted laser desorption/ionisation-time of flight and matrix-assisted laser desorption/ionisation-time of flight/time of flight-mass spectrometry. Proteomics 5, 2680–2688.

    Article  PubMed  CAS  Google Scholar 

  39. Umar, A., Luider, T. M., Foekens, J. A. and Pasa-Tolic, L. (2007) NanoLC-FT-ICR Ms improves proteome coverage attainable for approximately 3000 laser-microdissected breast carcinoma cells. Proteomics 7, 323–329.

    Article  PubMed  CAS  Google Scholar 

  40. Melle, C., Bogumil, R., Ernst, G., Schimmel, B., Bleul, A. and von Eggeling, F. (2006) Detection and identification of heat shock protein 10 as a biomarker in colorectal cancer by protein profiling. Proteomics 6, 2600–2608.

    Article  PubMed  CAS  Google Scholar 

  41. Melle, C., Ernst, G., Schimmel, B., Bleul, A., Koscielny, S., Wiesner, A., Bogumil, R., Moller, U., Osterloh, D., Halbhuber, K. J. and von Eggeling, F. (2003) Biomarker discovery and identification in laser microdissected head and neck squamous cell carcinoma with ProteinChip technology, two-dimensional gel electrophoresis, tandem mass spectrometry, and immunohistochemistry. Mol Cell Proteomics 2, 443–452.

    PubMed  CAS  Google Scholar 

  42. Zheng, Y., Xu, Y., Ye, B., Lei, J., Weinstein, M. H., O’Leary, M. P., Richie, J. P., Mok, S. C. and Liu, B. C. (2003) Prostate carcinoma tissue proteomics for biomarker discovery. Cancer 98, 2576–2582.

    Article  PubMed  Google Scholar 

  43. Cutler, P. (2003) Protein arrays: the current state-of-the-art. Proteomics 3, 3–18.

    Article  PubMed  CAS  Google Scholar 

  44. Dekker, L. J., Burgers, P. C., Guzel, C. and Luider, T. M. (2007) Ftms and TOF/TOF mass spectrometry in concert: identifying peptides with high reliability using matrix prespotted MALDI target plates. J Chromatogr B Analyt Technol Biomed Life Sci 847, 62–64.

    Article  PubMed  CAS  Google Scholar 

  45. Mustafa, D. A., Burgers, P. C., Dekker, L. J., Charif, H., Titulaer, M. K., Smitt, P. A., Luider, T. M. and Kros, J. M., (2007) Identification of glioma neovascularization-related proteins by using MALDI-FTMS and nano-LC fractionation to microdissected tumor vessels. Mol Cell Proteomics 6, 1147–1157.

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Josephine Nefkens Institute Erasmus Medical Center, Department of Pathology, Rotterdam, The Netherlands

    Dana Mustafa

  2. Josephine Nefkens Institute Erasmus Medical Center, Department of Pathology, Rotterdam, The Netherlands

    Johan M. Kros

  3. Josephine Nefkens Institute Erasmus Medical Center, Laboratories of Neuro-Oncology/Clinical and Cancer Proteomics, Rotterdam, The Netherlands

    Theo Luider

Authors
  1. Dana Mustafa
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Johan M. Kros
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Theo Luider
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Biomedical Research Foundation, Academy of Athens, Athens, Greece

    Antonia Vlahou

Rights and permissions

Reprints and permissions

Copyright information

© 2008 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Mustafa, D., Kros, J.M., Luider, T. (2008). Combining Laser Capture Microdissection and Proteomics Techniques. In: Vlahou, A. (eds) Clinical Proteomics. Methods in Molecular Biology™, vol 428. Humana Press. https://doi.org/10.1007/978-1-59745-117-8_9

Download citation

  • DOI: https://doi.org/10.1007/978-1-59745-117-8_9

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-837-9

  • Online ISBN: 978-1-59745-117-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation