Flow cytometric visualisation of cytokine production by CD3-CD56+ NK cells and CD3+CD56+ NK-T cells in whole blood.

Journal article

BACKGROUND: Natural killer (NK) cells produce multiple cytokines with potential immune regulatory roles. We standardised a whole-blood flow cytometry method to visualise intracellular cytokine production by NK cells for the study of NK cell biology and for clinical monitoring. METHODS: With a three-colour fluorescent labelling technique, specific cytokine production by NK or T cells was visualised directly in whole blood in the same sample after stimulation by phorbol 12-myristate 13-acetate (PMA) and ionomycin and by electronically gating on the CD3-ve/CD56+ve NK population or on the CD3+/CD56+ NK-T-cell population. RESULTS: Detectable levels of tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) but not of interleukin-2 (IL-2) or IL-4 were easily observed in NK cells. The visualisation of the cytokine production by NK cells was dependent on the addition of a Golgi transport inhibitor, Brefeldin A. Other known stimuli for NK cells (IL-2 and CD16 monoclonal antibody and incubation with K562, the NK-sensitive cell line) promoted IFN-gamma and TNF-alpha production in NK cells to a lesser extent than did PMA and ionomycin stimulation. CONCLUSIONS: This whole-blood flow cytometric assay appears to be an useful and easy method to examine cytokine production by NK cells and/or by CD3+CD56+ NK-T lymphocytes in patients with relevant diseases.

Authors: Mendes, R; Bromelow, K; Westby, M; Galea-Lauri, J; Smith, I

• Publication status: Published
• Journal: Cytometry
• Volume: 39
• Issue: 1
• Pages: 72-78
• Publication date: 2000-01-01
• DOI: 10.1002/(sici)1097-0320(20000101)39:1<72::aid-cyto10>3.0.co;2-r
• EISSN: 1097-0320
• ISSN: 0196-4763
• Language: English
• Keywords: Humans; K562 Cells; Protein Synthesis Inhibitors; Brefeldin A; Flow Cytometry; T-Lymphocytes; Monensin; Antigens, CD3; Killer Cells, Natural; Cytokines; Antigens, CD56