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. 1972 Jan;10(1):25–47.

Role of lymphocyte activation products (LAP) in cell-mediated immunity. I. Preparation and partial purification of guinea-pig LAP

D C Dumonde, D A Page, Margaret Matthew, R A Wolstencroft
PMCID: PMC1713095  PMID: 4111694

Abstract

Partial purification of soluble products of guinea-pig lymphocyte activation (LAP) was undertaken by fractional precipitation with ammonium sulphate, ion-exchange and Sephadex chromatography, and by immune precipitation of inducing antigen and of contaminating serum protein. During these purification steps the activity of macrophage migration-inhibition factor (MIF) was concentrated up to 1300-fold and separated from inducing antigen and serum protein. An endpoint assay was devised for expressing antigen-induced MIF activity of LAP fractions as weights of material giving 30% inhibition of migration (MI30 doses).

MIF activity precipitated between 50% and 80% saturated ammonium sulphate and eluted from DEAE-cellulose at pH 7·9 at intermediate salt concentrations (0·03–0·2 M phosphate). On Sephadex gel filtration MIF activity was concentrated in fractions of molecular weight range 56,000–82,000 with a smaller amount of activity eluting from 20,000–56,000. After immune precipitation of extraneous protein and elution from DEAE-cellulose, LAP material was found to have an MI30 dose of 0·4 μg.

Materials representative of antigen and serum protein-depleted MIF were selected for intralymphatic injection in order to determine whether MIF-rich LAP fractions were able to induce paracortical distension in guinea-pig lymph nodes (see following paper).

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Selected References

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

  1. Bennett B., Bloom B. R. Reactions in vivo and in vitro produced by a soluble substance associated with delayed-type hypersensitivity. Proc Natl Acad Sci U S A. 1968 Mar;59(3):756–762. doi: 10.1073/pnas.59.3.756. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bloom B. R., Bennett B. Mechanism of a reaction in vitro associated with delayed-type hypersensitivity. Science. 1966 Jul 1;153(3731):80–82. doi: 10.1126/science.153.3731.80. [DOI] [PubMed] [Google Scholar]
  3. David J. R. Delayed hypersensitivity in vitro: its mediation by cell-free substances formed by lymphoid cell-antigen interaction. Proc Natl Acad Sci U S A. 1966 Jul;56(1):72–77. doi: 10.1073/pnas.56.1.72. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Dumonde D. C., Maini R. N. The clinical significance of mediators of cellular immunity. Clin Allergy. 1971 Mar;1(1):123–139. doi: 10.1111/j.1365-2222.1971.tb02453.x. [DOI] [PubMed] [Google Scholar]
  5. Dumonde D. C., Wolstencroft R. A., Panayi G. S., Matthew M., Morley J., Howson W. T. "Lymphokines": non-antibody mediators of cellular immunity generated by lymphocyte activation. Nature. 1969 Oct 4;224(5214):38–42. doi: 10.1038/224038a0. [DOI] [PubMed] [Google Scholar]
  6. Kelly R. H., Wolstencroft R. A., Dumonde D. C., Balfour B. M. Role of lymphocyte activation products (LAP) in cell-mediated immunity. II. Effects of lymphocyte activation products on lymph node architecture and evidence for peripheral release of LAP following antigenic stimulation. Clin Exp Immunol. 1972 Jan;10(1):49–65. [PMC free article] [PubMed] [Google Scholar]
  7. McFARLANE A. S. Efficient trace-labelling of proteins with iodine. Nature. 1958 Jul 5;182(4627):53–53. doi: 10.1038/182053a0. [DOI] [PubMed] [Google Scholar]
  8. Möller G., Sjöberg O. Effect of antigenic competition on antigen-sensitive cells and on adoptively transferred immunocompetent cells. Cell Immunol. 1970 May;1(1):110–121. doi: 10.1016/0008-8749(70)90064-x. [DOI] [PubMed] [Google Scholar]
  9. Remold H. G., Katz A. B., Haber E., David J. R. Studies on migration inhibitory factor (MIF): recovery of MIF activity after purification by gel filtration and disc electrophoresis. Cell Immunol. 1970 May;1(1):133–145. doi: 10.1016/0008-8749(70)90066-3. [DOI] [PubMed] [Google Scholar]
  10. SCHEIDEGGER J. J. Une micro-méthode de l'immuno-electrophorèse. Int Arch Allergy Appl Immunol. 1955;7(2):103–110. [PubMed] [Google Scholar]
  11. Wolstencroft R. A., Dumonde D. C. In vitro studies of cell-mediated immunity. I. Induction of lymphocyte transformation by a soluble "mitogenic" factor derived from interaction of sensitized guinea-pig lymphoid cells with specific antigen. Immunology. 1970 Apr;18(4):599–610. [PMC free article] [PubMed] [Google Scholar]

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