Skip to main content
PMC home page
Immunology logoLink to Immunology
. 1987 Jan;60(1):1–5.

A novel approach to the identification of T-cell epitopes in Mycobacterium tuberculosis using human T-lymphocyte clones.

J R Lamb, D B Young
PMCID: PMC1453351  PMID: 2434413

Abstract

Current approaches to the analysis of antigens involved in the cellular immune response to mycobacterial infection rely on the initial identification and isolation of molecular components using monoclonal antibodies. In order to overcome the constraints of this approach, we have utilized a procedure involving T-cell recognition of antigens fractionated by polyacrylamide gel electrophoresis (SDS-PAGE) and added to proliferation assays after blotting onto nitrocellulose membranes. Analysis of human T-cell responses to Mycobacterium tuberculosis and Mycobacterium bovis BCG by this procedure revealed distinctive patterns of reactivity to different molecular weight components indicative of the selective recognition of immunodominant and species-specific determinants. Human T-cell clones were subsequently derived, and SDS-PAGE immunoblotting was used to identify the antigen recognized by each clone. Three epitopes defined by individual T-cell clones were identified on separate polypeptides with molecular weights 16,000-18,000 (clone P53), 18,000-20,000 (clone P57) and 52,000-55,000 (clone P35). This study demonstrates the potential application of T-cell cloning in conjunction with SDS-PAGE immunoblotting for the dissection and analysis of the cellular immune response to pathogenic agents during human infection.

Full text

PDF
1

Images in this article

3Figure 1
on p.3

Selected References

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

  1. Berzofsky J. A., Richman L. K., Killion D. J. Distinct H-2-linked Ir genes control both antibody and T cell responses to different determinants on the same antigen, myoglobin. Proc Natl Acad Sci U S A. 1979 Aug;76(8):4046–4050. doi: 10.1073/pnas.76.8.4046. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Emmrich F., Thole J., van Embden J., Kaufmann S. H. A recombinant 64 kilodalton protein of Mycobacterium bovis bacillus Calmette-Guerin specifically stimulates human T4 clones reactive to mycobacterial antigens. J Exp Med. 1986 Apr 1;163(4):1024–1029. doi: 10.1084/jem.163.4.1024. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Hahn H., Kaufmann S. H. The role of cell-mediated immunity in bacterial infections. Rev Infect Dis. 1981 Nov-Dec;3(6):1221–1250. doi: 10.1093/clinids/3.6.1221. [DOI] [PubMed] [Google Scholar]
  4. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  5. Lamb J. R., Green N. Analysis of the antigen specificity of influenza haemagglutinin-immune human T lymphocyte clones: identification of an immunodominant region for T cells. Immunology. 1983 Dec;50(4):659–666. [PMC free article] [PubMed] [Google Scholar]
  6. Results of a World Health Organization-sponsored workshop on monoclonal antibodies to Mycobacterium leprae. Infect Immun. 1985 May;48(2):603–605. doi: 10.1128/iai.48.2.603-605.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Results of a World Health Organization-sponsored workshop to characterize antigens recognized by mycobacterium-specific monoclonal antibodies. Infect Immun. 1986 Feb;51(2):718–720. doi: 10.1128/iai.51.2.718-720.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Thole J. E., Dauwerse H. G., Das P. K., Groothuis D. G., Schouls L. M., van Embden J. D. Cloning of Mycobacterium bovis BCG DNA and expression of antigens in Escherichia coli. Infect Immun. 1985 Dec;50(3):800–806. doi: 10.1128/iai.50.3.800-806.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Young D. B., Lamb J. R. T lymphocytes respond to solid-phase antigen: a novel approach to the molecular analysis of cellular immunity. Immunology. 1986 Oct;59(2):167–171. [PMC free article] [PubMed] [Google Scholar]
  10. Young D., Kent L., Rees A., Lamb J., Ivanyi J. Immunological activity of a 38-kilodalton protein purified from Mycobacterium tuberculosis. Infect Immun. 1986 Oct;54(1):177–183. doi: 10.1128/iai.54.1.177-183.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Young R. A., Bloom B. R., Grosskinsky C. M., Ivanyi J., Thomas D., Davis R. W. Dissection of Mycobacterium tuberculosis antigens using recombinant DNA. Proc Natl Acad Sci U S A. 1985 May;82(9):2583–2587. doi: 10.1073/pnas.82.9.2583. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Young R. A., Mehra V., Sweetser D., Buchanan T., Clark-Curtiss J., Davis R. W., Bloom B. R. Genes for the major protein antigens of the leprosy parasite Mycobacterium leprae. Nature. 1985 Aug 1;316(6027):450–452. doi: 10.1038/316450a0. [DOI] [PubMed] [Google Scholar]

Articles from Immunology are provided here courtesy of British Society for Immunology

RESOURCES