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Tolerizing Mice to Human Leukocytes: A Step Toward the Production of Monoclonal Antibodies Specific for Human Dendritic Cells

  • Una O’Doherty
  • William J. Swiggard
  • Yasunori Yamaguchi
  • Iris Kopeloff
  • Nina Bhardwaj
  • Ralph M. Steinman
  • Kayo Inaba
Chapter
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 329)

Abstract

Despite several attempts to isolate a mAb specific for human dendritic cells, none currently exists. Recent attempts have utilized an improved dendritic cell purification method to prepare immunogens1 and a rapid two-color flow cytometric screening procedure that allows large numbers of hybridoma supernatants to be examined in each fusion2. Yet these improvements have also failed, yielding only hybridomas that bind “shared” antigens expressed by both dendritic cells and other leukocytes. Dendritic cells express many shared antigens, including CD45 [leukocyte common antigen], CD40, leukocyte [β2] integrins CD11a and CD11c, CD54 [ICAM-1], CD44 [Pgp-1], CD58 [LFA-3], and the B7/BB1 antigen. Therefore, we are attempting to bias the immune response toward rarer, dendritic cell-specific clones by tolerizing or immunosuppressing our animals to shared antigens.

In one approach, adult mice held in barrier cages are injected with “nondendritic” cells and cyclophosphamide [CP], in order to ablate responding “nonspecific” B cell clones. Fifteen days after the last dose of CP, they are challenged with nondendritic cells. A week later they are bled, and serum antibody titers against nondendritic cells are determined by FACS, in order to demonstrate tolerance compared to controls injected with CP alone.

In the second approach, neonatal mice are injected with human T lymphoblasts at birth, followed by boosting at 1 week. In adulthood, they are challenged sequentially with sheep erythrocytes [sRBC], then with T blasts, to demonstrate that they can respond to unrelated cells but not to tolerogenic cells. One week after each kind of challenge, mice are bled and serum antibody levels are determined for treated and sham-injected mice.

When these two approaches were compared, CP led only to nonspecific immunosuppression, while neonatal injections produced selective, antigen-specific nonresponsiveness to the tolerizing T blasts.

Keywords

Dendritic Cell Sheep Erythrocyte Human Dendritic Cell Serum Antibody Level Shared Antigen 
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.

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References

  1. 1.
    P. S. Freudenthal and R. M. Steinman, The distinct surface of human blood dendritic cells, as observed after an improved isolation method, Proc. Natl. Acad. Sci. USA 87: 7698 (1990).PubMedCrossRefGoogle Scholar
  2. 2.
    J. P. Metlay, M. D. Witmer-Pack, R. Agger, M. T. Crowley, D. Lawless, and R. M. Steinman, The distinct leukocyte integrins of mouse spleen dendritic cells as identified with new hamster monoclonal antibodies, J. Exp. Med. 171: 1753 (1990).PubMedCrossRefGoogle Scholar
  3. 3.
    W. D. Matthew and P. H. Patterson, The production of a monoclonal antibody that blocks the action of a neurite outgrowth-promoting factor, Cold Spring Harbor Symp. Quant. Biol. 48: 625 (1983).PubMedCrossRefGoogle Scholar
  4. 4.
    S. Hockfield, A mAb to a unique cerebellar neuron generated by immunosuppression and rapid immunization, Science 237: 67 (1987).PubMedCrossRefGoogle Scholar
  5. 5.
    G. S. Golumbeski Jr. and R. L. Dimond, The use of tolerization in the production of monoclonal antibodies against minor antigenic determinants, Anal. Biochem. 154: 373 (1986).PubMedCrossRefGoogle Scholar
  6. 6.
    S. K. Ou, C. McDonald, and P. H. Patterson, Comparison of two techniques for targeting the production of monoclonal antibodies against particular antigens, J. Immunological Methods 145: 111 (1991).CrossRefGoogle Scholar
  7. 7.
    R. Schwartz and W. Dameshek, Drug-induced immunological tolerance, Nature 183: 1682 (1959).PubMedCrossRefGoogle Scholar
  8. 8.
    W. D. Matthew and A. W. Sandrock Jr., Cyclophosphamide treatment used to manipulate the immune response for the production of monoclonal antibodies, J. Immunological Methods 100: 73 (1987).CrossRefGoogle Scholar
  9. 9.
    C. V. Williams, C. L. Stechmann, and S. C. McLoon, Subtractive immunization techniques for the production of monoclonal antibodies to rare antigens, Biotechniques 12: 842 (1992).PubMedGoogle Scholar
  10. 10.
    M. C. Berenbaum, Immunology. A screen for agents inhibiting the immune response and the growth of tumours, Nature 196: 384 (1962).PubMedCrossRefGoogle Scholar
  11. 11.
    R. Hanan and J. Oyama, Inhibition of antibody formation in mature rabbits by contact with the antigen at an early age, J. Immunol. 73: 49 (1954).PubMedGoogle Scholar
  12. 12.
    F. J. Dixon and P. H. Maurer, Immunologic unresponsiveness induced by protein antigens, J. Exp. Med. 101: 245 (1954).CrossRefGoogle Scholar
  13. 13.
    R. T. Smith and R. A. Bridges, Immunological unresponsiveness in rabbits produced by neonatal injection of defined antigens, J. Exp. Med. 108: 227 (1958).PubMedCrossRefGoogle Scholar
  14. 14.
    R. E. Billingham, L. Brent, and P. B. Medawar, “Actively acquired tolerance” of foreign cells, Nature 172: 603 (1953).PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 1993

Authors and Affiliations

  • Una O’Doherty
    • 1
  • William J. Swiggard
    • 1
  • Yasunori Yamaguchi
    • 1
  • Iris Kopeloff
    • 1
  • Nina Bhardwaj
    • 1
  • Ralph M. Steinman
    • 1
  • Kayo Inaba
    • 2
  1. 1.Laboratory of Cellular Physiology and ImmunologyThe Rockefeller UniversityNew YorkUSA
  2. 2.Department of Zoology, Faculty of ScienceKyoto UniversityKyotoJapan

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