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Clinical and Experimental Immunology logoLink to Clinical and Experimental Immunology
. 1993 Apr;92(1):39–45. doi: 10.1111/j.1365-2249.1993.tb05945.x

Production and analysis of IgG monoclonal anti-DNA antibodies from systemic lupus erythematosus (SLE) patients.

M Ehrenstein 1, C Longhurst 1, D A Isenberg 1
PMCID: PMC1554857  PMID: 8385586

Abstract

This study compares recently devised methods for producing IgG anti-DNA MoAbs from patients with SLE and analyses the antibodies generated from one patient at different phases of disease. Lymphocytes from SLE patients were transformed with Epstein-Barr virus(EBV) and/or fused with a heteromyeloma cell line, CB-F7. Direct fusion with CB-F7 resulted in the highest proportion of IgG-secreting lines, whereas EBV transformation resulted in a high percentage of IgM-secreting lines. Using direct fusion, five IgM anti-DNA antibody-secreting hybridomas were generated using lymphocytes from a patient with relatively inactive SLE. Six months later when the disease was active, only IgG anti-DNA antibodies were produced. The antigen-binding patterns of the MoAbs were analysed. Only one of the IgM anti-DNA antibodies reacted with dsDNA by ELISA and none by Crithidia immunofluorescence, whereas two of the IgG antibodies reacted with dsDNA by ELISA and Crithidia but did not bind to ssDNA. Only the two IgG high affinity anti-dsDNA antibodies bound to histones, and this was enhanced by added DNA, whereas three IgM antibodies bound to cardiolipin. This study supports the notion that MoAbs derived from a patient with SLE represent those found in the serum of SLE patients at different stages of disease activity. The binding to histones by the two IgG anti-dsDNA antibodies supports the recently expressed view that antibodies binding DNA/histone may be important in the pathogenesis of SLE.

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Selected References

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  1. Bell D. A., Morrison B., VandenBygaart P. Immunogenic DNA-related factors. Nucleosomes spontaneously released from normal murine lymphoid cells stimulate proliferation and immunoglobulin synthesis of normal mouse lymphocytes. J Clin Invest. 1990 May;85(5):1487–1496. doi: 10.1172/JCI114595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brinkman K., Termaat R., Berden J. H., Smeenk R. J. Anti-DNA antibodies and lupus nephritis: the complexity of crossreactivity. Immunol Today. 1990 Jul;11(7):232–234. doi: 10.1016/0167-5699(90)90095-q. [DOI] [PubMed] [Google Scholar]
  3. Bron D., Delforge A., Lagneaux L., De Martynoff G., Bosmans E., Van der Auwera P., Snoeck R., Burny A., Stryckmans P. Production of human monoclonal IgG antibodies reacting with cytomegalovirus (CMV). J Immunol Methods. 1990 Jul 3;130(2):209–216. doi: 10.1016/0022-1759(90)90050-6. [DOI] [PubMed] [Google Scholar]
  4. Casali P., Notkins A. L. Probing the human B-cell repertoire with EBV: polyreactive antibodies and CD5+ B lymphocytes. Annu Rev Immunol. 1989;7:513–535. doi: 10.1146/annurev.iy.07.040189.002501. [DOI] [PubMed] [Google Scholar]
  5. Chan T. M., Frampton G., Staines N. A., Hobby P., Perry G. J., Cameron J. S. Different mechanisms by which anti-DNA MoAbs bind to human endothelial cells and glomerular mesangial cells. Clin Exp Immunol. 1992 Apr;88(1):68–74. doi: 10.1111/j.1365-2249.1992.tb03041.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dar O., Salaman M. R., Seifert M. H., Isenberg D. A. Spontaneous antibody-secreting cells against DNA and common environmental antigens in systemic lupus erythematosus. J Autoimmun. 1990 Oct;3(5):523–530. doi: 10.1016/s0896-8411(05)80018-6. [DOI] [PubMed] [Google Scholar]
  7. Ebling F., Hahn B. H. Restricted subpopulations of DNA antibodies in kidneys of mice with systemic lupus. Comparison of antibodies in serum and renal eluates. Arthritis Rheum. 1980 Apr;23(4):392–403. doi: 10.1002/art.1780230402. [DOI] [PubMed] [Google Scholar]
  8. Ehrenstein M. R., Leaker B., Isenberg D., Cambridge G. Production of human monoclonal antibodies to myeloperoxidase. Immunology. 1992 Dec;77(4):617–620. [PMC free article] [PubMed] [Google Scholar]
  9. Friguet B., Chaffotte A. F., Djavadi-Ohaniance L., Goldberg M. E. Measurements of the true affinity constant in solution of antigen-antibody complexes by enzyme-linked immunosorbent assay. J Immunol Methods. 1985 Mar 18;77(2):305–319. doi: 10.1016/0022-1759(85)90044-4. [DOI] [PubMed] [Google Scholar]
  10. Gladman D. D., Goldsmith C. H., Urowitz M. B., Bacon P., Bombardier C., Isenberg D., Kalunian K., Liang M. H., Maddison P., Nived O. Crosscultural validation and reliability of 3 disease activity indices in systemic lupus erythematosus. J Rheumatol. 1992 Apr;19(4):608–611. [PubMed] [Google Scholar]
  11. Grunow R., Jahn S., Porstmann T., Kiessig S. S., Steinkellner H., Steindl F., Mattanovich D., Gürtler L., Deinhardt F., Katinger H. The high efficiency, human B cell immortalizing heteromyeloma CB-F7. Production of human monoclonal antibodies to human immunodeficiency virus. J Immunol Methods. 1988 Feb 10;106(2):257–265. doi: 10.1016/0022-1759(88)90206-2. [DOI] [PubMed] [Google Scholar]
  12. Isenberg D. A., Collins C. Detection of cross-reactive anti-DNA antibody idiotypes on renal tissue-bound immunoglobulins from lupus patients. J Clin Invest. 1985 Jul;76(1):287–294. doi: 10.1172/JCI111959. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kalsi J. K., Isenberg D. A. Immortalisation of human antibody producing cells. Autoimmunity. 1992;13(3):249–258. doi: 10.3109/08916939209004831. [DOI] [PubMed] [Google Scholar]
  14. Klinman D. M., Shirai A., Ishigatsubo Y., Conover J., Steinberg A. D. Quantitation of IgM- and IgG-secreting B cells in the peripheral blood of patients with systemic lupus erythematosus. Arthritis Rheum. 1991 Nov;34(11):1404–1410. doi: 10.1002/art.1780341110. [DOI] [PubMed] [Google Scholar]
  15. MacGregor A. J., Kalsi J., Ravirajan C. T., Leaker B., Watts R., Winska-Wiloch H., Knight B., Norden A., Isenberg D. A., Cambridge G. Analysis of antibody reactivity in the sera of 42 patients with paraproteinaemia. Autoimmunity. 1992;13(2):101–105. doi: 10.3109/08916939209001910. [DOI] [PubMed] [Google Scholar]
  16. Manheimer-Lory A. J., Davidson A., Watkins D., Hannigan N. R., Diamond B. A. Generation and analysis of clonal IgM- and IgG-producing human B cell lines expressing an anti-DNA-associated idiotype. J Clin Invest. 1991 May;87(5):1519–1525. doi: 10.1172/JCI115162. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Olsson L., Kronstrøm H., Cambon-De Mouzon A., Honsik C., Brodin T., Jakobsen B. Antibody producing human-human hybridomas. I. Technical aspects. J Immunol Methods. 1983 Jun 24;61(1):17–32. doi: 10.1016/0022-1759(83)90004-2. [DOI] [PubMed] [Google Scholar]
  18. Ravirajan C. T., Kalsi J., Wiloch H. W., Barakat S., Tuaillon N., Irvine W., Cockayne A., Harris A., Williams D. G., Williams W. Antigen-binding diversity of human hybridoma autoantibodies derived from splenocytes of patients with SLE. Lupus. 1992 May;1(3):157–165. doi: 10.1177/096120339200100307. [DOI] [PubMed] [Google Scholar]
  19. Rumore P. M., Steinman C. R. Endogenous circulating DNA in systemic lupus erythematosus. Occurrence as multimeric complexes bound to histone. J Clin Invest. 1990 Jul;86(1):69–74. doi: 10.1172/JCI114716. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Sabbaga J., Pankewycz O. G., Lufft V., Schwartz R. S., Madaio M. P. Cross-reactivity distinguishes serum and nephritogenic anti-DNA antibodies in human lupus from their natural counterparts in normal serum. J Autoimmun. 1990 Apr;3(2):215–235. doi: 10.1016/0896-8411(90)90142-f. [DOI] [PubMed] [Google Scholar]
  21. Shoenfeld Y., Rauch J., Massicotte H., Datta S. K., André-Schwartz J., Stollar B. D., Schwartz R. S. Polyspecificity of monoclonal lupus autoantibodies produced by human-human hybridomas. N Engl J Med. 1983 Feb 24;308(8):414–420. doi: 10.1056/NEJM198302243080802. [DOI] [PubMed] [Google Scholar]
  22. Stollar B. D., Papalian M. Secondary structure in denatured DNA is responsible for its reaction with antinative DNA antibodies of systemic lupus erythematosus sera. J Clin Invest. 1980 Aug;66(2):210–219. doi: 10.1172/JCI109846. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Subiza J. L., Caturla A., Pascual-Salcedo D., Chamorro M. J., Gazapo E., Figueredo M. A., de la Concha E. G. DNA-anti-DNA complexes account for part of the antihistone activity found in patients with systemic lupus erythematosus. Arthritis Rheum. 1989 Apr;32(4):406–412. doi: 10.1002/anr.1780320409. [DOI] [PubMed] [Google Scholar]
  24. Symmons D. P., Coppock J. S., Bacon P. A., Bresnihan B., Isenberg D. A., Maddison P., McHugh N., Snaith M. L., Zoma A. S. Development and assessment of a computerized index of clinical disease activity in systemic lupus erythematosus. Members of the British Isles Lupus Assessment Group (BILAG). Q J Med. 1988 Nov;69(259):927–937. [PubMed] [Google Scholar]
  25. Tan E. M., Cohen A. S., Fries J. F., Masi A. T., McShane D. J., Rothfield N. F., Schaller J. G., Talal N., Winchester R. J. The 1982 revised criteria for the classification of systemic lupus erythematosus. Arthritis Rheum. 1982 Nov;25(11):1271–1277. doi: 10.1002/art.1780251101. [DOI] [PubMed] [Google Scholar]
  26. Watts R. A., Williams W., Le Page S., Norden A., Soltys A., Swana G., Addison I., Hay F. C., Isenberg D. A. Analysis of autoantibody reactivity and common idiotype PR4 expression of myeloma proteins. J Autoimmun. 1989 Oct;2(5):689–700. doi: 10.1016/s0896-8411(89)80007-1. [DOI] [PubMed] [Google Scholar]
  27. Watts R. A., Winska-Wiloch H., Muller S., Isenberg D. A. Analysis of factors affecting human hybridoma production. Hum Antibodies Hybridomas. 1990;1(3):160–165. [PubMed] [Google Scholar]
  28. Winkler T. H., Jahn S., Kalden J. R. IgG human monoclonal anti-DNA autoantibodies from patients with systemic lupus erythematosus. Clin Exp Immunol. 1991 Sep;85(3):379–385. doi: 10.1111/j.1365-2249.1991.tb05735.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. van Es J. H., Gmelig Meyling F. H., van de Akker W. R., Aanstoot H., Derksen R. H., Logtenberg T. Somatic mutations in the variable regions of a human IgG anti-double-stranded DNA autoantibody suggest a role for antigen in the induction of systemic lupus erythematosus. J Exp Med. 1991 Feb 1;173(2):461–470. doi: 10.1084/jem.173.2.461. [DOI] [PMC free article] [PubMed] [Google Scholar]

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