Multiple Color Immunofluorescence for Cytokine Detection at the Single Cell Level

Upon proper stimulation (e.g., crosslinking FcγRIIIA [CD16], interaction with antibody [Ab]-nonsensitized target cells, cytokines), mature natural killer (NK) cells produce and release cytokines involved in the modulation of immune responses (reviewed in refs. 1 ,2 ), including interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), GM-CSF, interleukin-5 (IL-5), and IL-10 (3 –7 ). Cytokine production in vitro is synergistically induced by combinations of these factors and by nonspecific stimuli, such as phorbol diesters and Ca²⁺ ionophores (5 ,8 ).