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Long-Term B-Lymphoid Cultures from Murine Bone Marrow Establishment and Cloning by Using Stromal Cell Line AC 6.21

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Basic Cell Culture Protocols

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 75))

Abstract

Nearly all hematopoietic cells in mammals derive from precursors that undergo much or all of their development in the bone marrow. In vitro models for many lineages are available and represent modifications of the original bone marrow culture system designed by Dexter and Lajtha (1). In this chapter, we describe a second bone marrow culture system, first reported in 1982 (2), that provides an in vitro environment selectively supporting long-term proliferation and differentiation of early B-lymphocyte lineage cells. This method can be used to obtain heterogeneous populations of immature precursors of the B-cell lineage greatly enriched from other hematopoietic cell types. Clonal populations can also be obtained by extension of this method to limiting dilution culture.

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References

  1. Dexter, T. M. and Lajtha, T. G. (1974) Proliferation of haemopoietic stem cells in vivo. Br. J. Haematol. 28, 525–530.

    Article  CAS  PubMed  Google Scholar 

  2. Whitlock, C. A. and Witte, O. N. (1982) Long-term culture of B lymphocytes and their precursors from murine bone marrow. Proc. Natl. Acad. Sci. USA 79, 3608–3612.

    Article  CAS  PubMed  Google Scholar 

  3. Coffman, R. L. and Weissman, I. L. (1983) Immunoglobulin gene rearrangement during pre-B cell differentiation. J. Mol. Cell. Immunol. 1, 31–38.

    CAS  PubMed  Google Scholar 

  4. Springer, T., Galfre, G., Secher, D. S., and Milstein, C. (1979) Mac-1: a macrophage differentiation antigen identified by monoclonal antibody. Eur. J. Immunol. 9, 301–306.

    Article  CAS  PubMed  Google Scholar 

  5. Holmes, K. L., Langdon, W. Y., Fredrickson, T. N., Coffman, R L., Hoffman, P. M., Hartley, J. W., and Morse, H. C. (1986) Analysis of neoplasmen induced by CAS-BR-M MuLV tumor extracts. J. Immunol. 137, 679–688.

    CAS  PubMed  Google Scholar 

  6. Ledbetter, J. A. and Herzenberg, L. A. (1979) Xenogeneic monoclonal antibodies to mouse lymphoid differentiation antigens. Immunol. Rev. 47, 63–90.

    Article  CAS  PubMed  Google Scholar 

  7. Lostrom, M. W., Stone, M. R., Tam, M., Burnette, W. N., Pinter, A., and Nowinski, R. C. (1979). Monoclonal antibodies against leukemia viruses: identification of six antigenic determinants on the p15(E) and gp70 envelope proteins. Virology 98, 336–350.

    Article  CAS  PubMed  Google Scholar 

  8. McGrath, M. S., Pillmer, E., and Weissman, I. L. (1980) Murine leukemogenesis. monoclonal antibodies to T cell determinants arrest T lymphoma cell proliferation. Nature 285, 259–261.

    Article  CAS  PubMed  Google Scholar 

  9. Dialynos, D. P., Wilde, D. B., Marrack, P., Pierrces, A., Wall, K. A., Havran, W., Otten, G., Loken, M. R., Pierres, M. R., Kappler, J., and Fitch, F. W. (1983) Characterization of the murine antigenic determinant, designated L3T4a, recognized by monoclonal antibody GK1.5: Expression of L3T4 by functional T cell clones appears to correlate primarily with class II MHC antigen restriction. Immunol. Rev. 74, 29–55.

    Article  Google Scholar 

  10. Hardy, R. (1984) Purification and coupling of fluorescent proteins for use in flow cytometry, in Handbook of Experimental Immunology, fourth ed. (Weir, D. M., Blackwell, C., and Herzenberg, L. A., eds.), Blackwell Scientific, Oxford, UK, pp. 146–155.

    Google Scholar 

  11. Whitlock, C. A., Tidmarsh, G. R., Muller-Sieburg, C. E., and Weissman, I. L. (1987) Bone marrow stromal cell lines with lymphopoietic activity express high levels of a pre-B neoplasia-associated molecule. Cell 48, 1009–1021.

    Article  CAS  PubMed  Google Scholar 

  12. Nottenberg, C. and Weissman, I. L. (1981) Cμ gene rearrangements of mouse immunoglobulin genes in normal B cells occurs on both expressed and nonexpressed chromosomes. Proc. Natl. Acad. Sci. USA 78, 484–488.

    Article  Google Scholar 

  13. Muller-Sieburg, C. E., Whitlock, C. A., and Weissman, I. L. (1986) Isolation of two early B lymphocyte progenitors from mouse marrow: a committed pre-pre-B cell and a clonogemc Thy-1lo hematopoietic stem cell. Cell 44, 653–662.

    Article  CAS  PubMed  Google Scholar 

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© 1997 Humana Press Inc., Totowa, NJ

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Whitlock, C.A., Muller-Sieburg, C.E. (1997). Long-Term B-Lymphoid Cultures from Murine Bone Marrow Establishment and Cloning by Using Stromal Cell Line AC 6.21. In: Pollard, J.W., Walker, J.M. (eds) Basic Cell Culture Protocols. Methods in Molecular Biology™, vol 75. Humana Press, Totowa, NJ. https://doi.org/10.1385/0-89603-441-0:231

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  • DOI: https://doi.org/10.1385/0-89603-441-0:231

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-0-89603-441-9

  • Online ISBN: 978-1-59259-561-7

  • eBook Packages: Springer Protocols

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