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Part of the book series: Methods in Molecular Medicine™ ((MIMM,volume 94))

Abstract

Two-dimensional electrophoresis results in an adequate resolution of the proteome of microorganisms to allow the detection and identification of specific antigens after blotting on membranes and overlaying the protein pattern with patient’s sera. The complement of all identified antigens presents the immunoproteome of a microorganism. All the antigens specific for a microorganism or even for a disease are identified by mass spectrometry. For identification, peptide mass fingerprinting is used, and post-translational modifications are detected by mass spectrometry MS/MS techniques. High-resolution two-dimensional electrophoresis and unambiguous identification are prerequisites for reliable results. After statistical analysis, the resulting antigens are candidates for diagnosis or vaccination and targets for therapy.

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References

  1. Smithies, O. and Poulik, M. D. (1956) Nature 177, 1033.

    Article  PubMed  CAS  Google Scholar 

  2. Kaltschmidt, E. and Wittmann, H. G. (1970) Ribosomal proteins. VII. Two-dimensional polyacrylamide gel electrophoresis for fingerprinting of ribosomal proteins. Anal. Biochem. 36, 401–412.

    Article  PubMed  CAS  Google Scholar 

  3. Klose, J. (1975) Protein mapping by combined isoelectric focusing and electrophoresis of mouse tissues. A novel approach to testing for induced point mutations in mammals. Humangenetik 26, 231–243.

    PubMed  CAS  Google Scholar 

  4. O’Farrell, P. H. (1975) High resolution two-dimensional electrophoresis of proteins. J. Biol. Chem. 250, 4007–4021.

    Google Scholar 

  5. Klose, J. and Kobalz, U. (1995) Two-dimensional electrophoresis of proteins: an updated protocol and implications for a functional analysis of the genome. Electrophoresis 16, 1034–1059.

    Article  PubMed  CAS  Google Scholar 

  6. Wasinger, V. C., Cordwell, S. J., Cerpa-Poljak, A., et al. (1995) Progress with gene-product mapping of the Mollicutes: Mycoplasma genitalium. Electrophoresis 16, 1090–1094.

    Article  PubMed  CAS  Google Scholar 

  7. Towbin, H., Staehelin, T., and Gordon, J. (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc. Natl. Acad. Sci. USA 76, 4350–4354.

    Article  PubMed  CAS  Google Scholar 

  8. Burnette, W. N. (1981) “Western blotting”: electrophoretic transfer of proteins from sodium dodecyl sulfate—polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal. Biochem. 112, 195–203.

    Article  PubMed  CAS  Google Scholar 

  9. Celis, J. E., Ratz, G. P., Madsen, P., et al. (1989) Computerized, comprehensive databases of cellular and secreted proteins from normal human embryonic lung MRC-5 fibroblasts: identification of transformation and/or proliferation sensitive proteins. Electrophoresis 10, 76–115.

    Article  PubMed  CAS  Google Scholar 

  10. Jungblut, P. R., Grabher, G., and Stoffler, G. (1999) Comprehensive detection of immunorelevant Borrelia garinii antigens by two-dimensional electrophoresis. Electrophoresis 20, 3611–3622.

    Article  PubMed  CAS  Google Scholar 

  11. Haas, G., Karaali, G., Ebermayer, K., et al. (2002) Immunoproteomics of Helicobacter pylori infection and relation to gastric disease. Proteomics 2, 313–324.

    Article  PubMed  CAS  Google Scholar 

  12. Jungblut, P., Thiede, B., Zimny-Arndt, U., et al. (1996) Resolution power of two-dimensional electrophoresis and identification of proteins from gels. Electrophoresis 17, 839–847.

    Article  PubMed  CAS  Google Scholar 

  13. Jungblut, P. R., Bumann, D., Haas, G., et al. (2000) Comparative proteome analysis of Helicobacter pylori. Mol. Microbiol. 36, 710–725.

    Article  PubMed  CAS  Google Scholar 

  14. Khyse-Andersen, J. (1984) Electroblotting of multiple gels: a simple apparatus without buffer tank for rapid transfer of proteins from polyacrylamide to nitrocellulose. J. Biochem. Biophys. Methods 10, 203–209.

    Article  Google Scholar 

  15. Jungblut, P., Eckerskorn, C., Lottspeich, F., and Klose, J. (1990) Blotting efficiency investigated by using two-dimensional electrophoresis, hydrophobic membranes and proteins from different sources. Electrophoresis 11, 581–588.

    Article  PubMed  CAS  Google Scholar 

  16. Krah, A., Miehlke, S., Pleissner, K. P., et al. (2003) Identification of candidate antigens for serologic detection of Helicobacter pylori infected patients with gastric carcinoma. Int. J. Cancer (in press).

    Google Scholar 

  17. Johannsson, K. E. (1986) Double replica electroblotting: a method to produce two replicas from gels. J. Biochem. Biophys. Methods 13, 197–203.

    Article  Google Scholar 

  18. Zeindl-Eberhart, E., Jungblut, P. R., and Rabes, H. M. (1997) A new method to assign immunodetected spots in the complex two-dimensional electrophoresis pattern. Electrophoresis 18, 799–801.

    Article  PubMed  CAS  Google Scholar 

  19. Pappin, D. J. (1997) Peptide mass fingerprinting using MALDI-TOF mass spectrometry. Methods Mol. Biol. 64, 165–173.

    PubMed  CAS  Google Scholar 

  20. Jungblut, P. R., Muller, E. C., Mattow, J., and Kaufmann, S. H. (2001) Proteomics reveals open reading frames in Mycobacterium tuberculosis H37Rv not predicted by genomics. Infect. Immun. 69, 5905–5907.

    Article  PubMed  CAS  Google Scholar 

  21. Jungblut, P. R. and Seifert, R. (1990) Analysis by high-resolution two-dimensional electrophoresis of differentiation-dependent alterations in cytosolic protein pattern of HL-60 leukemic cells. J. Biochem. Biophys. Methods 21, 47–58.

    Article  PubMed  CAS  Google Scholar 

  22. Doherty, N. S., Littman, B. H., Reilly, K., Swindell, A. C., Buss, J. M., and Anderson, N. L. (1998) Analysis of changes in acute-phase plasma proteins in an acute inflammatory response and in rheumatoid arthritis using two-dimensional gel electrophoresis. Electrophoresis 19, 355–363.

    Article  PubMed  CAS  Google Scholar 

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© 2004 Humana Press Inc.

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Krah, A., Jungblut, P.R. (2004). Immunoproteomics. In: Decler, J., Reischl, U. (eds) Molecular Diagnosis of Infectious Diseases. Methods in Molecular Medicine™, vol 94. Humana Press. https://doi.org/10.1385/1-59259-679-7:19

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  • DOI: https://doi.org/10.1385/1-59259-679-7:19

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-221-6

  • Online ISBN: 978-1-59259-679-9

  • eBook Packages: Springer Protocols

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