IL‐7 and immobilized Kit‐ligand stimulate serum‐ and stromal cell‐free cultures of precursor B‐cell lines and clones |
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| Authors: | Yohei Kawano Georg Petkau Ingrid Wolf Julia Tornack Fritz Melchers |
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| Affiliation: | 1. Department of Lymphocyte Development, Max‐Planck Institute for Infection Biology, Berlin, Germany;2. Deutsches Rheuma‐Forschungszentrum Berlin (DRFZ), Berlin, Germany |
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| Abstract: | Long‐term proliferating, DHJH‐rearranged mouse precursor B‐cell lines have previously been established in serum‐ and IL‐7‐containing media from fetal liver, but not from bone marrow. Serum and stromal cells expose these pre‐B cells to undefined factors, hampering accurate analyses of ligand‐dependent signaling, which controls pre‐B cell proliferation, survival, residence and migration. Here, we describe a novel serum‐free, stromal cell‐free culture system, which allows us to establish and maintain pre‐B cells not only from fetal liver, but also from bone marrow with practically identical efficiencies in proliferation, cloning and differentiation. Surprisingly, recombinant kit‐ligand, also called stem cell factor, produced as a kit‐ligand‐Fc fusion protein, suffices to replace stromal cells and serum, provided that it is presented to cultured pre‐B cells in an optimal density in plate‐bound, insolubilized, potentially crosslinking form. Additional recombinant CXCL12 and fibronectin have a minor influence on the establishment and maintenance of pre‐B cell lines and clones from fetal liver, but are necessary to establish such cell lines from bone marrow. |
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| Keywords: | IL‐7 Kit‐ligand Pre‐B cell Proliferation Serum‐free Stem cell factor Stromal cell‐free |
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