Intracellular Staining by Flow Cytometry
|Detection of IL‑12/IL‑23 p40 in Mature Dendritic Cells by Flow Cytometry. Mature dendritic cells were stained with Mouse Anti-Human IL‑12/IL‑23 p40 Fluorescein‑conjugated Monoclonal Antibody (Catalog # IC6092F, filled histogram) or isotype control antibody (Catalog # IC0041F, open histogram). To facilitate intracellular staining, cells were fixed with Flow Cytometry Fixation Buffer (Catalog # FC004) and permeabilized with Flow Cytometry Permeabilization/Wash Buffer I (Catalog # FC005). View our protocol for Staining Intracellular Molecules.|
Interleukin 23 (IL-23) is a heterodimeric cytokine composed of two disulfide-linked subunits, a p19 subunit that is unique to IL-23, and a p40 subunit that is shared with IL-12 (1‑5). The p19 subunit has homology to the p35 subunit of IL-12, as well as to other single chain cytokines such as IL-6 and IL-11. The p40 subunit is homologous to the extracellular domains of the hematopoietic cytokine receptors. Human p19 cDNA encodes a 189 amino acid residue (aa) precursor protein with a putative 19 aa signal peptide and 170 aa mature protein. Human and mouse p19 share 70% aa sequence identity. Although p19 is expressed by activated macrophages, dendritic cells, T cells, and endothelial cells, only activated macrophages and dendritic cells express p40 concurrently to produce IL-23. The functional IL-23 receptor complex consists of two receptor subunits, the IL-12 receptor beta 1 subunit (IL-12 R beta 1) and the IL-23-specific receptor subunit (IL-23 R). IL-23 has biological activities that are similar to, but distinct from IL-12. Both IL-12 and IL-23 induce proliferation and IFN-gamma production by human T cells. While IL-12 acts on both naïve and memory human T cells, the effects of IL-23 is restricted to memory T cells. In mouse, IL-23 but not IL-12, has also been shown to induce memory T cells to secret IL-17, a potent proinflammatory cytokine. IL-12 and IL-23 can induce IL-12 production from mouse splenic DC of both the CD8- and CD8+ subtypes, however only IL-23 can act directly on CD8+ DC to mediate immunogenic presentation of poorly immunogenic tumor/self peptide.
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