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. 1978 Mar 1;147(3):923–933. doi: 10.1084/jem.147.3.923

Complete replacement of serum by albumin, transferrin, and soybean lipid in cultures of lipopolysaccharide-reactive B lymphocytes

PMCID: PMC2184195  PMID: 305462

Abstract

Albumin, transferrin, and lipids can replace serum entirely for support of LPS-stimulated murine B lymphocytes in culture. In the presence of these compounds, growth and maturation to IgM and IgG secretion, induced by lipopolysaccharide (LPS), occurs at the same or higher efficiency in serum-free conditions as in conventional serum-containing medium, even at relatively low cell concentrations. In contrast to the rapid disappearance of LPS reactivity in conventional serum-containing medium, responsiveness remains at initial levels in serum-free conditions for 2 days before slowly declining. Overall lymphocyte survival is also markedly prolonged. In the presence of thymus "filler" cells, the serum-free conditions permit growth of every LPS-responsive cell to a clone of Ig-secreting cells at dilutions as low as a single reactive B cell per culture. The results have several important implications. These include the establishment for the first time of transferrin as a requirement for B lymphocyte responses in culture, and the availability now of conditions for the assay isolation of cell products regulating lymphocyte function, free of interference from undefined serum components.

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Selected References

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  1. Andersson J., Coutinho A., Lernhardt W., Melchers F. Clonal growth and maturation to immunoglobulin secretion in vitro of every growth-inducible B lymphocyte. Cell. 1977 Jan;10(1):27–34. doi: 10.1016/0092-8674(77)90136-2. [DOI] [PubMed] [Google Scholar]
  2. Andersson J., Coutinho A., Melchers F. Frequencies of mitogen-reactive B cells in the mouse. I. Distribution in different lymphoid organs from different inbred strains of mice at different ages. J Exp Med. 1977 Jun 1;145(6):1511–1519. doi: 10.1084/jem.145.6.1511. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Andersson J., Lafleur L., Melchers IgM in bone marrow-derived lymphocytes. Synthesis, surface deposition, turnover and carbohydrate composition in unstimulated mouse B cells. Eur J Immunol. 1974 Mar;4(3):170–180. doi: 10.1002/eji.1830040305. [DOI] [PubMed] [Google Scholar]
  4. Andersson J., Melchers F., Galanos C., Lüderitz O. The mitogenic effect of lipopolysaccharide on bone marrow-derived mouse lymphocytes. Lipid A as the mitogenic part of the molecule. J Exp Med. 1973 Apr 1;137(4):943–953. doi: 10.1084/jem.137.4.943. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Andersson J., Sjöberg O., Möller G. Induction of immunoglobulin and antibody synthesis in vitro by lipopolysaccharides. Eur J Immunol. 1972 Aug;2(4):349–353. doi: 10.1002/eji.1830020410. [DOI] [PubMed] [Google Scholar]
  6. Arai S., Yamane I., Tanno Y., Takishima T. Role of bovine serum albumin in blastoid transformation of lymphocytes by phytohemagglutinin. Proc Soc Exp Biol Med. 1977 Mar;154(3):444–448. doi: 10.3181/00379727-154-39690. [DOI] [PubMed] [Google Scholar]
  7. Bezkorovainy A., Zschocke R. H. Structure and function of transferrins. I. Physical, chemical, and iron-binding properties. Arzneimittelforschung. 1974 Apr;24(4):476–485. [PubMed] [Google Scholar]
  8. Broome J. D., Jeng M. W. Promotion of replication in lymphoid cells by specific thiols and disulfides in vitro. Effects on mouse lymphoma cells in comparison with splenic lymphocytes. J Exp Med. 1973 Sep 1;138(3):574–592. doi: 10.1084/jem.138.3.574. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chen R. F. Removal of fatty acids from serum albumin by charcoal treatment. J Biol Chem. 1967 Jan 25;242(2):173–181. [PubMed] [Google Scholar]
  10. Click R. E., Benck L., Alter B. J. Enhancement of antibody synthesis in vitro by mercaptoethanol. Cell Immunol. 1972 Jan;3(1):156–160. doi: 10.1016/0008-8749(72)90237-7. [DOI] [PubMed] [Google Scholar]
  11. Cooper R. A., Arner E. C., Wiley J. S., Shattil S. J. Modification of red cell membrane structure by cholesterol-rich lipid dispersions. A model for the primary spur cell defect. J Clin Invest. 1975 Jan;55(1):115–126. doi: 10.1172/JCI107901. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gronowicz E., Coutinho A., Melchers F. A plaque assay for all cells secreting Ig of a given type or class. Eur J Immunol. 1976 Aug;6(8):588–590. doi: 10.1002/eji.1830060812. [DOI] [PubMed] [Google Scholar]
  13. Hayashi I., Sato G. H. Replacement of serum by hormones permits growth of cells in a defined medium. Nature. 1976 Jan 15;259(5539):132–134. doi: 10.1038/259132a0. [DOI] [PubMed] [Google Scholar]
  14. Jonas A. Interaction of phosphatidylcholine with bovine serum albumin. Specificity and properties of the complexes. Biochim Biophys Acta. 1976 Mar 18;427(1):325–336. doi: 10.1016/0005-2795(76)90308-1. [DOI] [PubMed] [Google Scholar]
  15. Kincade P. W., Ralph P., Moore M. A. Growth of B-lymphocytes clones in semisolid culture is mitogen dependent. J Exp Med. 1976 May 1;143(5):1265–1270. doi: 10.1084/jem.143.5.1265. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. McKeehan W. L., Hamilton W. G., Ham R. G. Selenium is an essential trace nutrient for growth of WI-38 diploid human fibroblasts. Proc Natl Acad Sci U S A. 1976 Jun;73(6):2023–2027. doi: 10.1073/pnas.73.6.2023. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Melchers F., Andersson J. The kinectics of proliferation and maturation of mitogen-activated bone marrow-derived lymphocytes. Eur J Immunol. 1974 Oct;4(10):687–691. doi: 10.1002/eji.1830041010. [DOI] [PubMed] [Google Scholar]
  18. Melchers F., Coutinho A., Heinrich G., Andersson J. Continuous growth of mitogen-reactive B lymphocytes. Scand J Immunol. 1975;4(8):853–858. doi: 10.1111/j.1365-3083.1975.tb03728.x. [DOI] [PubMed] [Google Scholar]
  19. Metcalf D. Role of mercaptoethanol and endotoxin in stimulating B lymphocyte colony formation in vitro. J Immunol. 1976 Mar;116(3):635–638. [PubMed] [Google Scholar]
  20. Spieker-Polet H., Polet H. Identification of albumin as the serum factor essential for the growth of activated human lymphocytes. J Biol Chem. 1976 Feb 25;251(4):987–992. [PubMed] [Google Scholar]
  21. Stanley E. R., Palmer R. E., Sohn U. Development of methods for the quantitative in vitro analysis of androgen-dependent and autonomous Shionogi carcinoma 115 cells. Cell. 1977 Jan;10(1):35–44. doi: 10.1016/0092-8674(77)90137-4. [DOI] [PubMed] [Google Scholar]
  22. Takacs B. J., Rosenbusch J. P. Modification of Escherichia coli membranes in the prereplicative phase of phage T4 infection. Specificity of association and quantitation of bound phage proteins. J Biol Chem. 1975 Mar 25;250(6):2339–2350. [PubMed] [Google Scholar]
  23. Tormey D. C., Imrie R. C., Mueller G. C. Identification of transferrin as a lymphocyte growth promoter in human serum. Exp Cell Res. 1972 Sep;74(1):163–169. doi: 10.1016/0014-4827(72)90492-2. [DOI] [PubMed] [Google Scholar]
  24. Vogt A., Mishell R. I., Dutton R. W. Stimulation of DNA synthesis in cultures of mouse spleen cell suspensions by bovine transferrin. Exp Cell Res. 1969 Feb;54(2):195–200. doi: 10.1016/0014-4827(69)90232-8. [DOI] [PubMed] [Google Scholar]
  25. Zborowski J., Roerdink F., Scherphof G. Leakage of sucrose from phosphatidylcholine liposomes induced by interaction with serum albumin. Biochim Biophys Acta. 1977 Mar 29;497(1):183–191. doi: 10.1016/0304-4165(77)90151-9. [DOI] [PubMed] [Google Scholar]

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