Anti NFkB p65 (RelA), phospho Ser529  

TNF Induces phosphorylation of p65 in KBM-5 cells. Nuclear protein lysates prepared after 0, 5, 10, 15, 30 and 60 minutes of 0.1 nM TNF treatment of KBM-5 cells shows inducible phosphorylation using phospho specific polyclonal anti-human pS529 p65. Anti-beta-actin staining confirms loading of equivalent amounts of protein. HRP conjugated Gt-anti-Rabbit IgG was used to develop the blot using a chemiluminescent detection method. Other detection methods will yield similar results. Data contributed by Aggarwal BB, personal communication.

Anti-pS529 shows phospho p65 staining in carcinoma cells. western blot of total protein lysates from various human head and neck tumors shows phospho p65 staining in tumor cell lines using phospho specific polyclonal anti-human pS529 p65. Lanes 1-6 contain protein lysates from human squamal carcinoma cell lines. Lane 7 is a protein lysate from a primary culture of human keratinocytes. Lane 8 contains protein lysate from ATCC SCC9 cells (also a head and neck squamal carcinoma). Lane 9 contains lysate from EGF-induced human derived A431 cells. Lane 10 (not shown) contains a molecular weight standard. Concurrent staining with anti-beta microtubulin (not shown) was used to confirm equal protein loading in all lanes. HRP conjugated Gt-anti-Rabbit IgG was used to develop the blot using a chemiluminescent detection method. Other detection methods will yield similar results. Data contributed by Yu, M., NIH, personal communication.

Botanical extracts block the IL-1？？-induced phosphorylation and nuclear translocation of p65 in chondrocytes. Western blot analysis of IL-1？？-treated nuclear extracts. Serum-starved chondrocytes (0.1 √o 106 cells/mL) were pretreated with botanical extracts (10？Aa？og/ML each) for 4 hours followed by cotreatment with 10？Aang/mL IL-1？？ and botanical extracts for 1？Aah. Some chondrocyte cultures remained either untreated or were treated with 10？Aa？og/mL botanical extracts (each alone) or with 10？Aang/mL IL-1？？ alone for 1？Aah. Nuclear extracts were probed for phospho p65, (I) by western blot analysis using antibodies to p65, phospho-specific p65, and PARP (II, control). Treatment of chondrocytes with IL-1？？ (10？Aang/mL) revealed a clear increase in expression of phospho-p65 in the nuclear extracts (I). Co-treatment of chondrocytes with botanical extracts (all three) completely abolished the IL-1？？-dependent activation of phospho p65 in the nucleus (I). Synthesis of PARP remained unaffected in nuclear extracts (II). Data shown are representative of three independent experiments. Treatments: C (untreated control); IL-1？？; RC (Rosa canina); SA (Salix alba); UD (Urtica dioica). Figure provided by CiteAb. Source: Evid Based Complement Alternat Med, PMID: 22474508.