Skip to main content
PMC home page
The Journal of Experimental Medicine logoLink to The Journal of Experimental Medicine
. 1976 Jul 1;144(1):69–78. doi: 10.1084/jem.144.1.69

Antifluorescein affinity columns. Isolation and immunocompetence of lymphocytes that bind fluoresceinated antigens in vivo or in vitro

PMCID: PMC2190368  PMID: 1084410

Abstract

A new method for the isolation of specific immunocompetent lymphocytes has been described in which lymphocyte populations are exposed to fluoresceinated antigens (FLAGs) in vivo or in vitro, and the FLAG- binding cells retained on antifluorescein affinity columns. Specific cells are then eluted with an unrelated FL-labeled protein and shown to be fully immunocompetent. This methodology has been applied successfully in diverse antigenic systems including polymerized flagellin and TNP-specific B cells and alloantigen-reactive cytotoxic T lymphocytes. The method is rapid, inexpensive (requiring only antifluorescein beads), and can be applied to any antigens (or antibodies) in which a fluorescein group can be introduced.

Full Text

The Full Text of this article is available as a PDF (795.8 KB).

Selected References

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

  1. Aldo-Benson M., Borel Y. The tolerant cell: direct evidence for receptor blockade by tolerogen. J Immunol. 1974 May;112(5):1793–1803. [PubMed] [Google Scholar]
  2. Berke G., Sullivan K. A., Amos B. Rejection of ascites tumor allografts. I. Isolation, characterization, and in vitro reactivity of peritoneal lymphoid effector cells from BALB-c mice immune to EL4 leukosis. J Exp Med. 1972 Jun 1;135(6):1334–1350. doi: 10.1084/jem.135.6.1334. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Binz H., Wigzell H. Shared idiotypic determinants on B and T lymphocytes reactive against the same antigenic determinants. III. Physical fractionation of specific immunocompetent T lymphocytes by affinity chromatography using anti-idiotypic antibodies. J Exp Med. 1975 Nov 1;142(5):1231–1240. doi: 10.1084/jem.142.5.1231. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Choi T. K., Sleight D. R., Nisonoff A. General method for isolation and recovery of B cells bearing specific receptors. J Exp Med. 1974 Mar 1;139(3):761–766. doi: 10.1084/jem.139.3.761. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cunningham A. J., Szenberg A. Further improvements in the plaque technique for detecting single antibody-forming cells. Immunology. 1968 Apr;14(4):599–600. [PMC free article] [PubMed] [Google Scholar]
  6. Diener E. A new method for the enumeration of single antibody-producing cells. J Immunol. 1968 May;100(5):1062–1070. [PubMed] [Google Scholar]
  7. Diener E., Paetkau V. H. Antigen recognition: early surface-receptor phenomena induced by binding of a tritium-labeled antigen. Proc Natl Acad Sci U S A. 1972 Sep;69(9):2364–2368. doi: 10.1073/pnas.69.9.2364. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Edelman G. M., Rutishauser U., Millette C. F. Cell fractionation and arrangement on fibers, beads, and surfaces. Proc Natl Acad Sci U S A. 1971 Sep;68(9):2153–2157. doi: 10.1073/pnas.68.9.2153. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Elliott B. E., Haskill J. S., Axelrad M. A. Thymus-derived rosettes are not "helper" cells. J Exp Med. 1973 Nov 1;138(5):1133–1143. doi: 10.1084/jem.138.5.1133. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Greeley E., Berke G., Scott D. W. Receptors for antigen on lymphoid cells. I. Immunoadsorption of plaque-forming cells to poly-L-lysine-fixed antigen monolayers. J Immunol. 1974 Dec;113(6):1883–1890. [PubMed] [Google Scholar]
  11. Haas W., Layton J. E. Separation of antigen-specific lymphocytes. I. Enrichment of antigen-binding cells. J Exp Med. 1975 May 1;141(5):1004–1014. doi: 10.1084/jem.141.5.1004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Haas W. Separation of antigen-specific lymphocytes. II. Enrichment of hapten-specific antibody-forming cell precursors. J Exp Med. 1975 May 1;141(5):1015–1029. doi: 10.1084/jem.141.5.1015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hirsch J. S., Plescia O. J. Isolation of functional antigen-carrying spleen cells early in the course of the immune response of mice to sheep red blood cells. Immunol Commun. 1972;1(2):199–209. doi: 10.3109/08820137209022935. [DOI] [PubMed] [Google Scholar]
  14. Julius M. H., Masuda T., Herzenberg L. A. Demonstration that antigen-binding cells are precursors of antibody-producing cells after purification with a fluorescence-activated cell sorter. Proc Natl Acad Sci U S A. 1972 Jul;69(7):1934–1938. doi: 10.1073/pnas.69.7.1934. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Rittenberg M. B., Pratt K. L. Antitrinitrophenyl (TNP) plaque assay. Primary response of Balb/c mice to soluble and particulate immunogen. Proc Soc Exp Biol Med. 1969 Nov;132(2):575–581. doi: 10.3181/00379727-132-34264. [DOI] [PubMed] [Google Scholar]
  16. Rubin B. Regulation of helper cell activity by specifically adsorbable T lymphocytes. J Immunol. 1976 Jan;116(1):80–85. [PubMed] [Google Scholar]
  17. Schlossman S. F., Hudson L. Specific purification of lymphocyte populations on a digestible immunoabsorbent. J Immunol. 1973 Jan;110(1):313–319. [PubMed] [Google Scholar]
  18. Scott D. W. Cellular events in tolerance. V. Detection, isolation and fate of lymphoid cells which bind fluoresccinated antigen in vivo. Cell Immunol. 1976 Mar 15;22(2):311–322. doi: 10.1016/0008-8749(76)90032-0. [DOI] [PubMed] [Google Scholar]
  19. Shearer G. M. Cell-mediated cytotoxicity to trinitrophenyl-modified syngeneic lymphocytes. Eur J Immunol. 1974 Aug;4(8):527–533. doi: 10.1002/eji.1830040802. [DOI] [PubMed] [Google Scholar]
  20. Stulting R. D., Berke G. Nature of lymphocyte-tumor interaction. A general method for cellular immunoabsorption. J Exp Med. 1973 Apr 1;137(4):932–942. doi: 10.1084/jem.137.4.932. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Truffa-Bachi P., Wofsy L. Specific separation of cells on affinity columns. Proc Natl Acad Sci U S A. 1970 Jul;66(3):685–692. doi: 10.1073/pnas.66.3.685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wekerle H., Feldman M. T cell recognition of cell surface antigens. I. Specificity. Eur J Immunol. 1974 Apr;4(4):240–246. doi: 10.1002/eji.1830040403. [DOI] [PubMed] [Google Scholar]
  23. Wigzell H., Andersson B. Cell separation on antigen-coated columns. Elimination of high rate antibody-forming cells and immunological memory cells. J Exp Med. 1969 Jan 1;129(1):23–36. doi: 10.1084/jem.129.1.23. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Wigzell H., Mäkelä O. Separation of normal and immune lymphoid cells by antigen-coated coated columns. Antigen-binding characteristics of membrane antibodies as analyzed by hapten-protein antigens. J Exp Med. 1970 Jul 1;132(1):110–126. doi: 10.1084/jem.132.1.110. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The Journal of Experimental Medicine are provided here courtesy of The Rockefeller University Press

RESOURCES