Measurement of antigen-dependent interleukin-4 production by human peripheral blood mononuclear cells Introduction of an amplification step using ionomycin and phorbol myristate acetate
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The study of T cell responses in parasitic disease and allergy in humans has been limited by difficulties in the measurement of interleukin-4 (IL-4) in supernatants from antigen-stimulated peripheral blood mononuclear cells (PBMC). To obtain measurable amounts of IL-4 in vitro, we have added an amplification step to the antigen-specific response. Human PBMC were stimulated by tetanus toxoid (TT) or tuberculin (PPD) for 6 days and then pulsed with ionomycin and phorbol myristate acetate (PMA) for 24 h. TT-stimulated cells from nine revaccinated donors but not from seven unvaccinated donors and four that had only received childhood vaccinations against tetanus produced high levels of IL-4 (median (range) 1500 (300-3800), 316 (0-1600), and 270 (100-410) pg/ml, respectively, as measured by an enzyme linked immunosorbent assay, P = 0.005). PPD did not increase IL-4 production above the background level, although the majority of PPD-stimulated PBMC proliferated and produced interferon-γ (IFN-γ) in cultures without ionomycin and PMA. TT-induced IL-4 production correlated positively with proliferation. Culture supernatants did not interfere with IL-4 immunoreactivity and failed to affect ionomycin and PMA induced IL-4 production. The findings suggest that proliferating antigen-specific T cells were the source of IL-4 in these experiments. The method should prove useful for comparing the IL-4 producing ability of antigen-specific T cells from different individuals.
Originalsprog | Engelsk |
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Tidsskrift | Journal of Immunological Methods |
Vol/bind | 156 |
Udgave nummer | 2 |
Sider (fra-til) | 239-245 |
Antal sider | 7 |
ISSN | 0022-1759 |
DOI | |
Status | Udgivet - 8 dec. 1992 |
Bibliografisk note
Funding Information:
The work was supported by the Danish Biotechnology program (Center for Signal Peptides), the Danish Medical Research Council (grant 12-9796-2), the Danish Research Council for Developmental Research (grant 104.Dan.8/ 357), and by the Commission of the European Community (grant TS2-0059-DK\[AM\]).
ID: 328570442