Intracellular Staining by Flow Cytometry
|Detection of BAFF/BLyS/TNFSF13B in Mouse Splenocytes by Flow Cytometry. Mouse splenocytes were stained with Rat Anti-Mouse BAFF/BLyS/TNFSF13B Alexa Fluor® 488‑conjugated Monoclonal Antibody (Catalog # IC1357G, filled histogram) or isotype control antibody (Catalog # IC006G, 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.|
BAFF (also known as TALL-1, BLyS, THANK) is a type II transmembrane glycoprotein belonging to the TNF superfamily and has been designated as TNF superfamily member 13B (TNFSF13B). Mouse BAFF is a 309 amino acid (aa) protein consisting of a 248 aa extracellular domain, a 21 aa transmembrane region and a 45 aa cytoplasmic tail (1, 2). BAFF has the typical structural characteristics of the TNF superfamily ligands. It is a homotrimeric protein having the structurally conserved motif known as TNF homology domain (3, 4). A higher ordered structure composed of a cluster of trimeric units resembling the structure of a viral capsid has also been reported (4). Mouse BAFF may be shed from the cell surface by proteolytic cleavage between R126 and Ala 127 to yield a soluble form of the protein detectable in serum (1, 5). Within the TNF superfamily BAFF shares the highest homology (48%) with APRIL (1). BAFF shares with APRIL the ability to bind to BCMA and TACI and also binds specifically to BAFF receptor(BAFF R, also known as BR3 or TNFSFR13C), which is the principal BAFF receptor (6-8). All three receptors are type III transmembrane proteins that are expressed in B cells. BAFF and APRIL can form active heteromers that bind TACI (9). BAFF is expressed in peripheral blood mononuclear cells, in spleen and lymph nodes. Its expression in resting monocytes is upregulated by IFN-alpha, IFN-beta, LPS and IL-10. BAFF provides critical survival signals to a subset of B cells with intermediate maturation status (T2 B cells) during the immune response (10). BAFF also plays an important role in the development of lymphoid tissue and enhances the survival of activated memory B cells (7, 11). Human and mouse BAFF share 86% aa sequence identity (1).
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