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. 1977 Jun;28(3):526–534.

Colony formation by human T lymphocytes in agar medium.

M H Claësson, M B Rodger, G R Johnson, S Whittingham, D Metcalf
PMCID: PMC1541018  PMID: 330061

Abstract

An improved method is described for growing human T-lymphocyte colonies in agar medium containing phytohaemagglutinin (PHA) and sheep red blood cells (SRBC). Cluster and colony growth was obtained when blood mononuclear cells were plated directly in the agar-medium (one-step procedure) or after incubation of cells in liquid medium with PHA (two-step procedure). In the one-step procedure approximately 1 per 100 cells plated formed a cluster containing four to fifty cells. In the two-step procedure 1 per 20 cells plated formed a cluster or a colony (more than fifty cells). The proliferating cells were shown to be sheep-erythrocyte rosette-forming cells (E-RFC). Optimal proliferation was dependent on the presence of phagocytic cells in the cell suspensions cultured. No growth occurred in cultures depleted of E-RFC. Detailed studies of the cycle, velocity sedimentation, and density of the cells plated showed that the majority of cluster- and colony-forming cells were small non-cycline lymphocytes with a sedimentation velocity of 4 mm/hr, and a density between 1-069 and 1-077 g/cm3.

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Selected References

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

  1. Bradley T. R., Metcalf D. The growth of mouse bone marrow cells in vitro. Aust J Exp Biol Med Sci. 1966 Jun;44(3):287–299. doi: 10.1038/icb.1966.28. [DOI] [PubMed] [Google Scholar]
  2. Brostoff J. A simple technique for counting rosettes using acridine orange. J Immunol Methods. 1974 Aug;5(3):303–303. doi: 10.1016/0022-1759(74)90115-x. [DOI] [PubMed] [Google Scholar]
  3. Dwyer J. M., Mackay I. R. Antigen-binding lymphocytes in human blood. Lancet. 1970 Jan 24;1(7639):164–167. doi: 10.1016/s0140-6736(70)90406-x. [DOI] [PubMed] [Google Scholar]
  4. Fibach E., Gerassi E., Sachs L. Induction of colony formation in vitro by human lymphocytes. Nature. 1976 Jan 15;259(5539):127–129. doi: 10.1038/259127a0. [DOI] [PubMed] [Google Scholar]
  5. Kaplan M. E., Clark C. An improved rosetting assay for detection of human T lymphocytes. J Immunol Methods. 1974 Jul;5(2):131–135. doi: 10.1016/0022-1759(74)90003-9. [DOI] [PubMed] [Google Scholar]
  6. Levis W. R., Robbins J. H. Effect of glass-adherent cells on the blastogenic response of 'purified' lymphocytes to phytohemagglutinin. Exp Cell Res. 1970 Jul;61(1):153–158. doi: 10.1016/0014-4827(70)90269-7. [DOI] [PubMed] [Google Scholar]
  7. Lundgren G., Zukoski C. F., Möller G. Differential effects of human granulocytes and lymphocytes on human fibroblasts in vitro. Clin Exp Immunol. 1968 Oct;3(8):817–836. [PMC free article] [PubMed] [Google Scholar]
  8. Metcalf D., Nossal G. J., Warner N. L., Miller J. F., Mandel T. E., Layton J. E., Gutman G. A. Growth of B-lymphocyte colonies in vitro. J Exp Med. 1975 Dec 1;142(6):1534–1549. doi: 10.1084/jem.142.6.1534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Miller R. G., Phillips R. A. Separation of cells by velocity sedimentation. J Cell Physiol. 1969 Jun;73(3):191–201. doi: 10.1002/jcp.1040730305. [DOI] [PubMed] [Google Scholar]
  10. Moore M. A., Williams N. Analysis of proliferation and differentiation of foetal granulocyte-macrophage progenitor cells in haemopoietic tissue. Cell Tissue Kinet. 1973 Sep;6(5):461–476. doi: 10.1111/j.1365-2184.1973.tb01634.x. [DOI] [PubMed] [Google Scholar]
  11. Moretta L., Ferrarini M., Durante M. L., Mingari M. C. Expression of a receptor for IgM by human T cells in vitro. Eur J Immunol. 1975 Aug;5(8):565–569. doi: 10.1002/eji.1830050812. [DOI] [PubMed] [Google Scholar]
  12. Peters J. H. Contact cooperation in stimulated lymphocytes. I. Influence of cell contact on unspecifically stimulated lymphocytes. Exp Cell Res. 1972 Sep;74(1):179–186. doi: 10.1016/0014-4827(72)90495-8. [DOI] [PubMed] [Google Scholar]
  13. Rozenszajn L. A., Shoham D., Kalechman I. Clonal proliferation of PHA-stimulated human lymphocytes in soft agar culture. Immunology. 1975 Dec;29(6):1041–1055. [PMC free article] [PubMed] [Google Scholar]
  14. Sinclair W. K. Hydroxyurea: effects on Chinese hamster cells grown in culture. Cancer Res. 1967 Feb;27(2):297–308. [PubMed] [Google Scholar]
  15. Wilson J. D., Dalton G. Human T lymphocyte colonies in agar: a comparison with other T cell assays in healthy subjects and cancer patients. Aust J Exp Biol Med Sci. 1976 Feb;54(1):27–34. doi: 10.1038/icb.1976.3. [DOI] [PubMed] [Google Scholar]

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