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. 1985 Nov;82(21):7414–7418. doi: 10.1073/pnas.82.21.7414

Enrichment of hematopoietic precursor cells and cloning of multipotential B-lymphocyte precursors.

J P McKearn, J McCubrey, B Fagg
PMCID: PMC391355  PMID: 3933007

Abstract

A simple one-step isolation technique significantly enriched mouse fetal liver cells that respond to interleukin 3 (IL-3), a multilineage hematopoietic growth factor. The fetal liver cell subpopulation isolated with monoclonal antibody AA4 contained 50- to 100-fold higher frequencies of multipotential (CFU-mix) or restricted (CFU-G/M, BFU-E) erythroid/myeloid precursors as well as precursors that differentiate to become mature B lymphocytes [CFU-mix = erythroid and myeloid colony-forming unit(s); CFU-G/M = CFU-granulocyte/macrophage; BFU-E = burst-forming unit-erythroid]. The B-lymphocyte precursors could be cloned in single-cell cultures when IL-3-containing supernatants were present. Growth of these clones was supported by purified IL-3 but not by purified IL-2. Stable growth has been maintained for greater than 6 mo in the presence of IL-3. Such clones express on their cell surface low amounts of class I major histocompatibility complex antigens and high amounts of AA4, GF1, and leukocyte common glycoprotein 200 antigens. They lack detectable rearrangements of their Ig-encoding genes [joining region heavy and light (kappa, lambda) chain genes], even after subcloning, but maintain their capacity to differentiate to mature B lymphocytes committed to multiple Ig specificities.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Blomberg B., Traunecker A., Eisen H., Tonegawa S. Organization of four mouse lambda light chain immunoglobulin genes. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3765–3769. doi: 10.1073/pnas.78.6.3765. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Clark-Lewis I., Schrader J. W. P cell-stimulating factor: biochemical characterization of a new T cell-derived factor. J Immunol. 1981 Nov;127(5):1941–1947. [PubMed] [Google Scholar]
  3. Dexter T. M., Garland J., Scott D., Scolnick E., Metcalf D. Growth of factor-dependent hemopoietic precursor cell lines. J Exp Med. 1980 Oct 1;152(4):1036–1047. doi: 10.1084/jem.152.4.1036. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Gillis S., Ferm M. M., Ou W., Smith K. A. T cell growth factor: parameters of production and a quantitative microassay for activity. J Immunol. 1978 Jun;120(6):2027–2032. [PubMed] [Google Scholar]
  5. Gillis S., Mochizuki D. Y., Conlon P. J., Hefeneider S. H., Ramthun C. A., Gillis A. E., Frank M. B., Henney C. S., Watson J. D. Molecular characterization of interleukin 2. Immunol Rev. 1982;63:167–209. doi: 10.1111/j.1600-065x.1982.tb00415.x. [DOI] [PubMed] [Google Scholar]
  6. Hapel A. J., Lee J. C., Farrar W. L., Ihle J. N. Establishment of continuous cultures of thy1.2+, Lyt1+, 2-T cells with purified interleukin 3. Cell. 1981 Jul;25(1):179–186. doi: 10.1016/0092-8674(81)90242-7. [DOI] [PubMed] [Google Scholar]
  7. Ihle J. N., Keller J., Greenberger J. S., Henderson L., Yetter R. A., Morse H. C., 3rd Phenotypic characteristics of cell lines requiring interleukin 3 for growth. J Immunol. 1982 Oct;129(4):1377–1383. [PubMed] [Google Scholar]
  8. Ihle J. N., Keller J., Henderson L., Klein F., Palaszynski E. Procedures for the purification of interleukin 3 to homogeneity. J Immunol. 1982 Dec;129(6):2431–2436. [PubMed] [Google Scholar]
  9. Ihle J. N., Keller J., Oroszlan S., Henderson L. E., Copeland T. D., Fitch F., Prystowsky M. B., Goldwasser E., Schrader J. W., Palaszynski E. Biologic properties of homogeneous interleukin 3. I. Demonstration of WEHI-3 growth factor activity, mast cell growth factor activity, p cell-stimulating factor activity, colony-stimulating factor activity, and histamine-producing cell-stimulating factor activity. J Immunol. 1983 Jul;131(1):282–287. [PubMed] [Google Scholar]
  10. Iscove N. N., Roitsch C. A., Williams N., Guilbert L. J. Molecules stimulating early red cell, granulocyte, macrophage, and megakaryocyte precursors in culture: similarity in size, hydrophobicity, and charge. J Cell Physiol Suppl. 1982;1:65–78. doi: 10.1002/jcp.1041130412. [DOI] [PubMed] [Google Scholar]
  11. Kincade P. W., Lee G., Scheid M. P., Blum M. D. Characterization of murine colony-forming B cells. II. Limits to in vitro maturation, Lyb-2 expression, resolution of IgD+ subsets, and further population analysis. J Immunol. 1980 Feb;124(2):947–953. [PubMed] [Google Scholar]
  12. 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]
  13. Marcu K. B., Banerji J., Penncavage N. A., Lang R., Arnheim N. 5' flanking region of immunoglobulin heavy chain constant region genes displays length heterogeneity in germlines of inbred mouse strains. Cell. 1980 Nov;22(1 Pt 1):187–196. doi: 10.1016/0092-8674(80)90167-1. [DOI] [PubMed] [Google Scholar]
  14. McCubrey J., Risser R. Activation of nonexpressed endogenous ecotropic murine leukemia virus by transfection of genomic DNA into embryo cells. J Virol. 1983 Mar;45(3):950–955. doi: 10.1128/jvi.45.3.950-955.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. McKearn J. P., Baum C., Davie J. M. Cell surface antigens expressed by subsets of pre-B cells and B cells. J Immunol. 1984 Jan;132(1):332–339. [PubMed] [Google Scholar]
  16. McKearn J. P., Rosenberg N. Mapping cell surface antigens on mouse pre-B cell lines. Eur J Immunol. 1985 Mar;15(3):295–298. doi: 10.1002/eji.1830150316. [DOI] [PubMed] [Google Scholar]
  17. Quintáns J., Lefkovits I. Precursor cells specific to sheep red cells in nude mice. Estimation of frequency in the microculture system. Eur J Immunol. 1973 Jul;3(7):392–397. doi: 10.1002/eji.1830030704. [DOI] [PubMed] [Google Scholar]
  18. Smilek D. E., Boyd H. C., Wilson D. B., Zmijewski C. M., Fitch F. W., McKearn T. J. Monoclonal rat anti-major histocompatibility complex antibodies display specificity for rat, mouse, and human target cells. J Exp Med. 1980 May 1;151(5):1139–1150. doi: 10.1084/jem.151.5.1139. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  20. Springer T., Galfrè G., Secher D. S., Milstein C. Monoclonal xenogeneic antibodies to murine cell surface antigens: identification of novel leukocyte differentiation antigens. Eur J Immunol. 1978 Aug;8(8):539–551. doi: 10.1002/eji.1830080802. [DOI] [PubMed] [Google Scholar]
  21. Stanley E. R. Colony-stimulating factor (CSF) radioimmunoassay: detection of a CSF subclass stimulating macrophage production. Proc Natl Acad Sci U S A. 1979 Jun;76(6):2969–2973. doi: 10.1073/pnas.76.6.2969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Williams N., Eger R. R., Jackson H. M., Nelson D. J. Two-factor requirement for murine megakaryocyte colony formation. J Cell Physiol. 1982 Jan;110(1):101–104. doi: 10.1002/jcp.1041100116. [DOI] [PubMed] [Google Scholar]

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