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Immunochemical Measurement of Complement Components and Activation Products

  • Reinhard Würzner
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 150)

Abstract

Activation of the complement system by the classical, mannan binding lectic (MBL) or alternative pathway results in the generation of multiple complement proteolytic cleavage products, recruited from these inactive precursor molecules in a sequentially proceeding cascade ( Chapter 1). This leads to an alteration of their antigenic pattern and thus to the consumption of the nativemolecule. Concomitantly, complement activation initiates formation of multi-molecular complexes such as the C3 or C5 convertases or the terminal complement complex (TCC). The latter can be generated on biological membranes or in the fluid phase (C5b-9). Activation-dependent changes on molecules or complexes can be revealed by the disappearance of native-restricted epitopes, and by the appearance of neoepitopes (1), and are usually assessed by means of monoclonal antibodies in enzyme-linked immunosorbent acids (ELISAs). This chapter focuses on the sensitive and specific quantitation of (native) complement proteins to evaluate the inactivated (background) status, and also on that of their fragments and complexes to reliably assess complement activation in vivo, with particular respect to neoepitopes of C9 appearing only when C9 is incorporated into the terminal complement complex.

Keywords

Complement Activation Complement Protein Membrane Attack Complex Coating Buffer Immunochemical Assay 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

References

  1. 1.
    Mollnes T. E. and Harboe M. (1993) Neoepitope expression during complement activation—a model for detecting antigenic changes in proteins and activation of cascades. Immunologist 1, 43–49.Google Scholar
  2. 2.
    Whaley K. (1985) Methods in Complement for Clinical Immunologists. Churchill-Livingstone, Edinburgh.Google Scholar
  3. 3.
    Harrison R. A. (1996) Purification, assay, and characterization of complement proteins from plasma, in Handbook of Experimental Immunology (Herzenberg L. A., Weir D. M., Herzenberg L. A., and Blackwell C., eds.), Blackwell, Cambridge, MA, pp. 75.1–75.50.Google Scholar
  4. 4.
    Kirschfink M. (1997) The clinical laboratory: testing the complement system, in The Complement System (Rother K., Till G. O., and Hänsch G. M., eds.), Springer, Berlin, pp. 522–547.Google Scholar
  5. 5.
    Whaley K. and North J. (1997) Haemolytic assays for whole complement activity and individual components, in Complement: A Practical Approach (Dodds A. W. and Sim R. B., eds.), IRL Press, Oxford, pp. 19–47.Google Scholar
  6. 6.
    Oppermann M., Höpken U., and Gätze O. (1992) Assessment of complement activation in vivo. Immunopharmacology 24, 119–134.PubMedCrossRefGoogle Scholar
  7. 7.
    Würzner R., Mollnes T. E., and Morgan B. P. (1997) Immunochemical assays for complement components, in Immunochemistry 2: A Practical Approach (Johnstone A. P. and Turner M. W., eds.), Oxford University Press, Oxford, pp. 197–223.Google Scholar
  8. 8.
    Würzner R. (1993) Monoclonal antibodies against the terminal complement components, in Activators and Inhibitors of Complement (Sim R. B., ed.), Kluwer Academic, Doordrecht, The Netherlands, pp. 167–180.Google Scholar
  9. 9.
    Falk R. J., Dalmasso A. P., Kim Y., Tsai C. H., Scheinman J. I., Gewurz H., and Michael A. F. (1983) Neoantigen of the polymerized ninth component of complement. Characterization of a monoclonal antibody and immunohistochemi-cal localization in renal disease. J. Clin. Invest. 72, 560–573.PubMedCrossRefGoogle Scholar
  10. 10.
    Mollnes T. E., Lea T., Harboe M., and Tschopp J. (1985) Monoclonal antibodies recognizing a neoantigen of poly(C9) detect the human terminal complement complex in tissue and plasma. Scand. J. Immunol. 22, 183–195.Google Scholar
  11. 11.
    Hugo F., Jenne D., and Bhakdi S. (1985) Monoclonal antibodies against neoantigens of the terminal C5b-9 complex of human complement. Biosci. Rep. 5, 649–658.PubMedCrossRefGoogle Scholar
  12. 12.
    Kolb W. P., Morrow P. R., Jensen F. C., and Tamerius J. D. (1988) Development of a highly sensitive capture EIA for the quantification of the SC5b-9 complex in human plasma using a monoclonal antibody reactive with a poly-C9 neoantigenic determinant. Complement 5, 213–214.Google Scholar
  13. 13.
    Kusunoki Y., Takekoshi Y., and Nagasawa S. (1990) Using polymerized C9 to produce a monoclonal antibody against a neoantigen of the human terminal complement complex. J. Pharmacobiodyn. 13, 454–460.PubMedGoogle Scholar
  14. 14.
    Würzner R., Schulze M., Happe L., Franzke A., Bieber F. A., Oppermann M., and Götze O. (1991) Inhibition of terminal complement complex formation and cell lysis by monoclonal antibodies. Compl. Inflamm. 8, 328–340.Google Scholar
  15. 15.
    Würzner R., Xu H., Franzke A., Schulze M., Peters J. H., and Götze O. (1991) Blood dendritic cells carry terminal complement complexes on their cell surface as detected by newly developed neoepitope specific monoclonal antibodies. Immunology 74, 132–138.PubMedGoogle Scholar
  16. 16.
    Kemp P. A., Spragg J. H., Brown J. C., Morgan B. P., Gunn C. A., and Taylor P. W. (1992) Immunohistochemical determination of complement activation in joint tissues of patients with rheumatoid arthritis and osteoarthritis using neoantigen-specific monoclonal antibodies. J. Clin. Lab. Immunol. 37, 147–162.PubMedGoogle Scholar
  17. 17.
    Tschopp J. and Mollnes T. E. (1986) Antigenic crossreactivity of the alpha subunit of complement component C8 with the cysteine-rich domain shared by complement component C9 and low density lipoprotein receptor. Proc. Natl. Acad. Sci. USA 83, 4223–4227.PubMedCrossRefGoogle Scholar
  18. 18.
    Mollnes T. E., Lea T., Froland S. S., and Harboe M. (1985) Quantification of the terminal complement complex in human plasma by an enzyme-linked immunosorbent assay based on monoclonal antibodies against a neoantigen of the complex. Scand. J. Immunol. 22, 197–202.PubMedCrossRefGoogle Scholar
  19. 19.
    Hugo F., Krämer S., and Bhakdi S. (1987) Sensitive ELISA for quantitating the terminal membrane C5b-9 and fluid phase SC5b-9 complex of human complement. J. Immunol. Meth. 99, 243–251.CrossRefGoogle Scholar
  20. 20.
    Mollnes T. E., Garred P., and Bergseth G. (1988) Effect of time, temperature and anticoagulants on in vitro complement activation: consequences for collection and preservation of samples to be examined for complement activation. Clin. Exp. Immunol. 73, 484–488.PubMedGoogle Scholar
  21. 21.
    Würzner R., Orren A., Potter P., Morgan B. P., Ponard D., Späth P., et al. (1991) Functionally active complement proteins C6 and C7 detected in C6-or C7-deficient individuals. Clin. Exp. Immunol., 83, 430–437.PubMedCrossRefGoogle Scholar
  22. 22.
    Würzner R., Platonov A. E., Beloborodov V. B., Pereverzev A. I., Vershinina I. V., Fernie B. A., et al. (1996) How partial C7 deficiency with chronic and recurrent bacterial infections can mimic total C7 deficiency: temporary restoration of host C7 level following plasma transfusion. Immunology 88, 407–411.PubMedCrossRefGoogle Scholar
  23. 23.
    Witzel-Schlömp K., Hobart M. J., Fernie B. A., Orren A., Würzner R., Rittner C., et al. (1998) Heterogeneity in the genetic basis of human complement C9 deficiency. Immunogenetics 48, 144–147.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 2000

Authors and Affiliations

  • Reinhard Würzner
    • 1
  1. 1.Institut für HygieneInnsbruck UniversityInnsbruckAustria

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