Intracellular Staining by Flow Cytometry
|Detection of IL‑10 in Human PBMCs by Flow Cytometry. Human peripheral blood mononuclear cells (PBMCs) either (A) stimulated to induce TH2 cells or (B) unstimulated were stained with Rabbit Anti-Human IL‑10 Alexa Fluor® 488‑conjugated Monoclonal Antibody (Catalog # IC9210G) and Mouse Anti-Human CD4 Alexa Fluor® 700‑conjugated Monoclonal Antibody (Catalog # FAB3791N). Quadrant markers were set based on control antibody staining (Catalog # IC1051G). To facilitate intracellular staining, cells were fixed with paraformaldehyde and permeabilized with methanol. View our protocol for Staining Intracellular Molecules.|
Interleukin 10, also known as cytokine synthesis inhibitory factor (CSIF), is the charter member of the IL-10 family of alpha -helical cytokines that also includes IL-19, IL‑20, IL-22, IL-24, and IL-26/AK155 (1, 2). IL-10 is secreted by many activated hematopoietic cell types as well as hepatic stellate cells, keratinocytes, and placental cytotrophoblasts (2‑5). Mature human IL-10 shares 72%‑86% amino acid sequence identity with bovine, canine, equine, feline, mouse, ovine, porcine, and rat IL-10. Whereas human IL-10 is active on mouse cells, mouse IL-10 does not act on human cells (6, 7). IL-10 is a 178 amino acid molecule that contains two intrachain disulfide bridges and is expressed as a 36 kDa noncovalently associated homodimer (6, 8, 9). The IL-10 dimer binds to two IL-10 R alpha /IL-10 R1 chains, resulting in recruitment of two IL-10 R beta /IL-10 R2 chains and activation of a signaling cascade involving JAK1, TYK2, and STAT3 (10). IL-10 R beta does not bind IL-10 by itself but is required for signal transduction (1). IL-10 R beta also associates with IL‑20 R alpha, IL-22 R alpha, or IL-28 R alpha to form the receptor complexes for IL-22, IL-26, IL-28, and IL‑29 (11‑13). IL-10 is a critical molecule in the control of viral infections and allergic and autoimmune inflammation (14‑16). It promotes phagocytic uptake and Th2 responses but suppresses antigen presentation and Th1 proinflammatory responses (2).
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