Skip to main content
NCBI home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1985 Oct;76(4):1418–1426. doi: 10.1172/JCI112119

Monoclonal antibodies against complement 3 neoantigens for detection of immune complexes and complement activation. Relationship between immune complex levels, state of C3, and numbers of receptors for C3b.

M T Aguado, J D Lambris, G C Tsokos, R Burger, D Bitter-Suermann, J D Tamerius, F J Dixon, A N Theofilopoulos
PMCID: PMC424092  PMID: 2932466

Abstract

C3-bearing immune complexes and C3 activation products were detected by using two monoclonal antibodies, one specific for a neoantigenic determinant on C3c and the other for C3d. To quantitate immune complexes, the anti-C3c or anti-C3d antibodies were fixed to microtiter plates and reacted with test plasma. The binding of C3-bearing immune complexes in this plasma was then measured with radioisotope- or enzyme-labeled anti-human IgG. To test for C3 breakdown products, solid-phase monoclonal antibody to the C3d neoantigen was reacted with EDTA-plasma samples, and fixed iC3b or C3d was measured with a polyclonal anti-C3 antibody. Patients with autoimmune diseases, such as systemic lupus erythematosus, rheumatoid arthritis, and Sjogren's syndrome, and paracoccidioidomycosis were found to contain immune complexes bearing C3b/iC3b or C3d. In most conditions, there were more C3d-containing immune complexes than C3b/iC3b. Although CR1 (C3b receptors) rapidly converted immune complex-bound iC3b to C3dg/C3d and lupus patients had reduced CR1, no correlation between the state of C3 on circulating immune complexes or levels of immune complexes and CR1 numbers was seen. However, levels of C3-fixing ICs correlated with levels of C3 activation products. This assay system with monoclonal antibodies to neoantigens expressed on activated, but not native, C3 provides sensitive and specific means for detecting and classifying C3-fixing immune complexes and for assessing C3 activation.

Full text

PDF
1418

Images in this article

1423Image
on p.1423

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Altman A., Cardenas J. M., Houghten R. A., Dixon F. J., Theofilopoulos A. N. Antibodies of predetermined specificity against chemically synthesized peptides of human interleukin 2. Proc Natl Acad Sci U S A. 1984 Apr;81(7):2176–2180. doi: 10.1073/pnas.81.7.2176. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Anderson C. L., Stillman W. S. Raji cell assay for immune complexes. Evidence for detection of Raji-directed immunoglobulin G antibody in sera from patients with systemic lupus erythematosus. J Clin Invest. 1980 Aug;66(2):353–360. doi: 10.1172/JCI109863. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Arend W. P., Teller D. C., Mannik M. Molecular composition and sedimentation characteristics of soluble antigen-antibody complexes. Biochemistry. 1972 Oct 24;11(22):4063–4072. doi: 10.1021/bi00772a008. [DOI] [PubMed] [Google Scholar]
  4. Arvin A. M., Miller J. J., 3rd Acid labile alpha-interferon in sera and synovial fluids from patients with juvenile arthritis. Arthritis Rheum. 1984 May;27(5):582–585. doi: 10.1002/art.1780270517. [DOI] [PubMed] [Google Scholar]
  5. Berkowicz A., Rosenkvist J., Sørensen H. A single step immunoelectrophoresis method for the quantitation of complement C3c in biological fluids. J Immunol Methods. 1983 Jun 10;60(3):277–288. doi: 10.1016/0022-1759(83)90286-7. [DOI] [PubMed] [Google Scholar]
  6. Burger R., Deubel U., Hadding U., Bitter-Suermann D. Identification of functionally relevant determinants on the complement component C3 with monoclonal antibodies. J Immunol. 1982 Nov;129(5):2042–2050. [PubMed] [Google Scholar]
  7. Chaplin H., Monroe M. C., Lachmann P. J. Further studies of the C3g component of the alpha 2D fragment of human C3. Clin Exp Immunol. 1983 Mar;51(3):639–646. [PMC free article] [PubMed] [Google Scholar]
  8. Dobson N. J., Lambris J. D., Ross G. D. Characteristics of isolated erythrocyte complement receptor type one (CR1, C4b-C3b receptor) and CR1-specific antibodies. J Immunol. 1981 Feb;126(2):693–698. [PubMed] [Google Scholar]
  9. Dykman T. R., Hatch J. A., Atkinson J. P. Polymorphism of the human C3b/C4b receptor. Identification of a third allele and analysis of receptor phenotypes in families and patients with systemic lupus erythematosus. J Exp Med. 1984 Mar 1;159(3):691–703. doi: 10.1084/jem.159.3.691. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Eisenberg R. A., Theofilopoulos A. N., Dixon F. J. Use of bovine conglutinin for the assay of immune complexes. J Immunol. 1977 Apr;118(4):1428–1434. [PubMed] [Google Scholar]
  11. Fearon D. T. Regulation of the amplification C3 convertase of human complement by an inhibitory protein isolated from human erythrocyte membrane. Proc Natl Acad Sci U S A. 1979 Nov;76(11):5867–5871. doi: 10.1073/pnas.76.11.5867. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fearon D. T., Wong W. W. Complement ligand-receptor interactions that mediate biological responses. Annu Rev Immunol. 1983;1:243–271. doi: 10.1146/annurev.iy.01.040183.001331. [DOI] [PubMed] [Google Scholar]
  13. Friguet B., Djavadi-Ohaniance L., Goldberg M. E. Some monoclonal antibodies raised with a native protein bind preferentially to the denatured antigen. Mol Immunol. 1984 Jul;21(7):673–677. doi: 10.1016/0161-5890(84)90053-1. [DOI] [PubMed] [Google Scholar]
  14. Hansen O., Jensenius J. C., Folkersen J., Brandslund I. Production of antibody against C3c epitopes which are inaccessible on native C3. J Immunol Methods. 1983 Jul 15;61(2):245–252. doi: 10.1016/0022-1759(83)90168-0. [DOI] [PubMed] [Google Scholar]
  15. Hay F. C., Nineham L. J., Roitt I. M. Routine assay for the detection of immune complexes of known immunoglobulin class using solid phase C1q. Clin Exp Immunol. 1976 Jun;24(3):396–400. [PMC free article] [PubMed] [Google Scholar]
  16. Iida K., Mornaghi R., Nussenzweig V. Complement receptor (CR1) deficiency in erythrocytes from patients with systemic lupus erythematosus. J Exp Med. 1982 May 1;155(5):1427–1438. doi: 10.1084/jem.155.5.1427. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Iida K., Nussenzweig V. Complement receptor is an inhibitor of the complement cascade. J Exp Med. 1981 May 1;153(5):1138–1150. doi: 10.1084/jem.153.5.1138. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Inada Y., Kamiyama M., Kanemitsu T., Hyman C. L., Clark W. S. Studies on immune adherence (C3b) receptor activity of human erythrocytes: relationship between receptor activity and presence of immune complexes in serum. Clin Exp Immunol. 1982 Oct;50(1):189–197. [PMC free article] [PubMed] [Google Scholar]
  19. Jacobs R. J., Reichlin M. Generation of low m.w., C3-bearing immunoglobulin in human serum. J Immunol. 1983 Jun;130(6):2775–2781. [PubMed] [Google Scholar]
  20. Lachmann P. J., Oldroyd R. G., Milstein C., Wright B. W. Three rat monoclonal antibodies to human C3. Immunology. 1980 Nov;41(3):503–515. [PMC free article] [PubMed] [Google Scholar]
  21. Lachmann P. J., Pangburn M. K., Oldroyd R. G. Breakdown of C3 after complement activation. Identification of a new fragment C3g, using monoclonal antibodies. J Exp Med. 1982 Jul 1;156(1):205–216. doi: 10.1084/jem.156.1.205. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Lachmann P. J., Voak D., Oldroyd R. G., Downie D. M., Bevan P. C. Use of monoclonal anti-C3 antibodies to characterise the fragments of C3 that are found on erythrocytes. Vox Sang. 1983;45(5):367–372. doi: 10.1111/j.1423-0410.1983.tb01928.x. [DOI] [PubMed] [Google Scholar]
  23. Lambris J. D., Alsenz J., Schulz T. F., Dierich M. P. Mapping of the properdin-binding site in the third component of complement. Biochem J. 1984 Jan 1;217(1):323–326. doi: 10.1042/bj2170323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Law S. K., Lichtenberg N. A., Levine R. P. Evidence for an ester linkage between the labile binding site of C3b and receptive surfaces. J Immunol. 1979 Sep;123(3):1388–1394. [PubMed] [Google Scholar]
  25. McConahey P. J., Dixon F. J. A method of trace iodination of proteins for immunologic studies. Int Arch Allergy Appl Immunol. 1966;29(2):185–189. doi: 10.1159/000229699. [DOI] [PubMed] [Google Scholar]
  26. Medof M. E., Iida K., Mold C., Nussenzweig V. Unique role of the complement receptor CR1 in the degradation of C3b associated with immune complexes. J Exp Med. 1982 Dec 1;156(6):1739–1754. doi: 10.1084/jem.156.6.1739. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Medof M. E., Prince G. M., Mold C. Release of soluble immune complexes from immune adherence receptors on human erythrocytes is mediated by C3b inactivator independently of Beta 1H and is accompanied by generation of C3c. Proc Natl Acad Sci U S A. 1982 Aug;79(16):5047–5051. doi: 10.1073/pnas.79.16.5047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Miyakawa Y., Yamada A., Kosaka K., Tsuda F., Kosugi E., Mayumi M. Defective immune-adherence (C3b) receptor on erythrocytes from patients with systemic lupus erythematosus. Lancet. 1981 Sep 5;2(8245):493–497. doi: 10.1016/s0140-6736(81)90882-5. [DOI] [PubMed] [Google Scholar]
  29. Molenaar J. L., ten Velde A., Pondman K. W. Anti-C3a, anti-C3B (anti-B) and their reaction with the anaphylatoxic fragment C3a. J Immunol. 1973 Mar;110(3):702–708. [PubMed] [Google Scholar]
  30. NISONOFF A., WISSLER F. C., LIPMAN L. N., WOERNLEY D. L. Separation of univalent fragments from the bivalent rabbit antibody molecule by reduction of disulfide bonds. Arch Biochem Biophys. 1960 Aug;89:230–244. doi: 10.1016/0003-9861(60)90049-7. [DOI] [PubMed] [Google Scholar]
  31. Niemann M. A., Kearney J. F., Volanakis J. E. The use of monoclonal antibodies as probes of the three-dimensional structure of human complement factor D. J Immunol. 1984 Feb;132(2):809–815. [PubMed] [Google Scholar]
  32. Nydegger U. E., Zubler R. H., Gabay R., Joliat G., Karagevrekis C. H., Lambert P. H., Miescher P. A. Circulating complement breakdown products in patients with rheumatoid arthritis. Correlation between plasma C3d, circulating immune complexes, and clinical activity. J Clin Invest. 1977 May;59(5):862–868. doi: 10.1172/JCI108708. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Olds L. C., Miller J. J., 3rd Anti-F(ab')2 antibodies that interfere with interpretation of the anti-C3 assay for immune complexes in children with rheumatic diseases. Arthritis Rheum. 1984 Mar;27(3):330–336. doi: 10.1002/art.1780270313. [DOI] [PubMed] [Google Scholar]
  34. Pangburn M. K. Activation of complement via the alternative pathway. Fed Proc. 1983 Jan;42(1):139–143. [PubMed] [Google Scholar]
  35. Pereira A. B., Theofilopoulos A. N., Dixon F. J. Detection and partial characterization of circulating immune complexes with solid-phase anti-C3. J Immunol. 1980 Aug;125(2):763–770. [PubMed] [Google Scholar]
  36. Perrin L. H., Lambert P. H., Miescher P. A. Complement breakdown products in plasma from patients with systemic lupus erythematosus and patients with membranoproliferative or other glomerulonephritis. J Clin Invest. 1975 Jul;56(1):165–176. doi: 10.1172/JCI108065. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Ross G. D., Lambris J. D., Cain J. A., Newman S. L. Generation of three different fragments of bound C3 with purified factor I or serum. I. Requirements for factor H vs CR1 cofactor activity. J Immunol. 1982 Nov;129(5):2051–2060. [PubMed] [Google Scholar]
  38. Ross G. D. Structure and function of membrane complement receptors. Summary. Fed Proc. 1982 Dec;41(14):3089–3093. [PubMed] [Google Scholar]
  39. Salacinski P. R., McLean C., Sykes J. E., Clement-Jones V. V., Lowry P. J. Iodination of proteins, glycoproteins, and peptides using a solid-phase oxidizing agent, 1,3,4,6-tetrachloro-3 alpha,6 alpha-diphenyl glycoluril (Iodogen). Anal Biochem. 1981 Oct;117(1):136–146. doi: 10.1016/0003-2697(81)90703-x. [DOI] [PubMed] [Google Scholar]
  40. Tamerius J. D., Pangburn M. K., Müller-Eberhard H. J. Selective inhibition of functional sites of cell-bound C3b by hybridoma-derived antibodies. J Immunol. 1982 Jan;128(1):512–514. [PubMed] [Google Scholar]
  41. Theofilopoulos A. N., Dixon F. J. Detection of immune complexes: techniques and implications. Hosp Pract. 1980 May;15(5):107-11, 114-7, 121. doi: 10.1080/21548331.1980.11946602. [DOI] [PubMed] [Google Scholar]
  42. Theofilopoulos A. N., Dixon F. J. The biology and detection of immune complexes. Adv Immunol. 1979;28:89–220. doi: 10.1016/s0065-2776(08)60800-7. [DOI] [PubMed] [Google Scholar]
  43. Theofilopoulos A. N., Wilson C. B., Dixon F. J. The Raji cell radioimmune assay for detecting immune complexes in human sera. J Clin Invest. 1976 Jan;57(1):169–182. doi: 10.1172/JCI108257. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. West C., Davis N. C., Forristal J., Herbst J., Spitzer R. Antigenic determinants of human beta-1c and beta-1g-globulins. J Immunol. 1966 Apr;96(4):650–658. [PubMed] [Google Scholar]
  45. Wilson J. G., Wong W. W., Schur P. H., Fearon D. T. Mode of inheritance of decreased C3b receptors on erythrocytes of patients with systemic lupus erythematosus. N Engl J Med. 1982 Oct 14;307(16):981–986. doi: 10.1056/NEJM198210143071604. [DOI] [PubMed] [Google Scholar]
  46. Zubler R. H., Lange G., Lambert P. H., Miescher P. A. Detection of immune complexes in unheated sera by modified 125I-Clq binding test. Effect of heating on the binding of Clq by immune complexes and application of the test to systemic lupus erythematosus. J Immunol. 1976 Jan;116(1):232–235. [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES