Detection of IFN‑ gamma in Human PBMCs by Flow Cytometry.
Human peripheral blood mononuclear cells (PBMCs) treated with 50 ng/mL PMA, 1 μg/mL Ionomycin, and 3 μM Monensin overnight were stained with Mouse Anti-Human CD3 epsilon APC‑conjugated Monoclonal Antibody (Catalog # FAB100A) and either (A) Mouse Anti-Human IFN‑ gamma Alexa Fluor® 488‑conjugated Monoclonal Antibody (Catalog # IC285G) or (B) Mouse IgG2B Alexa Fluor 488 Isotype Control (Catalog # IC0041G). To facilitate intracellular staining, cells were fixed and permeabilized with FlowX FoxP3 Fixation & Permeabilization Buffer Kit (Catalog # FC012). View our protocol for Staining Intracellular Molecules.
Preparation and Storage
The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Protect from light. Do not freeze.
12 months from date of receipt, 2 to 8 °C as supplied.
Interferon-gamma (IFN-gamma ), also known as type II or immune interferon, exerts a wide range of immunoregulatory activities and is considered to be the prototype proinflammatory cytokine (1, 2). Mature human IFN-gamma exists as a non-covalently linked homodimer of 20-25 kDa variably glycosylated subunits (3). It shares 90% amino acid (aa) sequence identity with rhesus IFN-gamma, 59-64% with bovine, canine, equine, feline, and porcine IFN‑ gamma, and 37-43% with cotton rat, mouse, and rat IFN-gamma. IFN-gamma dimers bind to IFN-gamma RI (alpha subunits) which then interact with IFN-gamma RII (beta subunits) to form the functional receptor complex of two alpha and two beta subunits. Inclusion of IFN-gamma RII increases the binding affinity for ligand and the efficiency of signal transduction (4, 5). IFN-gamma is produced by a variety of immune cells under inflammatory conditions, notably by T cells and NK cells (6). It plays a key role in host defense by promoting the development and activation of Th1 cells, chemoattraction and activation of monocytes and macrophages, upregulation of antigen presentation molecules, and immunoglobulin class switching in B cells. It also exhibits antiviral, antiproliferative, and apoptotic effects (6, 7). In addition, IFN-gamma functions as an anti-inflammatory mediator by promoting the development of regulatory T cells and inhibiting Th17 cell differentiation (8, 9). The pleiotropic effects of IFN-gamma contribute to the development of multiple aspects of atherosclerosis (7).
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Marsters, S.A. et al. (1995) Proc. Natl. Acad. Sci. USA 92:5401.
Krause, C.D. et al. (2000) J. Biol. Chem. 275:22995.
Schroder, K. et al. (2004) J. Leukoc. Biol. 75:163.
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