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Disease-specific ex vivo stimulation of whole blood for cytokine production: applications in the study of tuberculosis.

Authors
Type
Published Article
Journal
Journal of Immunological Methods
0022-1759
Publisher
Elsevier
Publication Date
Volume
222
Issue
1-2
Pages
145–153
Identifiers
PMID: 10022381
Source
Medline
License
Unknown

Abstract

As a simple method for field studies to assess the cytokine-status of patients with tuberculosis (TB), the use of whole blood instead of isolated cells has advantages, especially since the risk of contamination is minimal. Therefore, cytokine production in whole blood cultures was determined using non-specific and disease-specific stimuli. Heparinized blood from healthy volunteers was either incubated in closed vacutainer tubes or in tissue culture wells after dilution in culture medium. Dose-response and kinetics were investigated for the production of TNFalpha, IL-1beta, IL-1ra, IL-10 and IFNgamma. Patients with TB and healthy individuals were examined for IFN-gamma production in whole blood. In the absence of a stimulus, the production of cytokines is negligible in whole blood cultures. LPS induces the production of TNFalpha, IL-1beta, IL-1ra and IL-10; PHA induces the production of IFNgamma and IL-10. Live BCG induces the production of proinflammatory cytokines, irrespective of tuberculin skin status. In contrast, PPD and MTB-culture filtrate induce production of IFNgamma in skin-test positive and not in skin-test negative healthy subjects. Five out of 13 patients with TB had a low antigen-specific IFNgamma production, suggestive of a minimal or absent specific T-cell response. For most purposes, cultures in closed vacutainer tubes are optimal. If one wishes to focus on T-cell cytokines or if only small volumes of blood are available, dilution of whole blood in culture medium before incubation in tissue culture wells may be preferable.

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