Western blotting was performed as previously described (Chen et al., 2002 (link); Xiao et al., 2019 (link)). Antibodies anti-myeloid differentiation primary response gene 88 (MyD88) (D80F5; #4283), anti-phospho-IKKα/β (Ser176/180) (16A6; #2697), anti-IKKα (#2682), Phospho-IKKα/β (Ser176/180) (16A6, #2697), anti-IκBα (44D4,#4812), anti-Phospho-IκBα (Ser32/36) (5A5, #9246), anti-NF-κB p65 (D14E12, #8242) and anti-Phospho-NF-κB p65 (Ser536) (93H1, #3033) were purchased from Cell Signaling Technology.
Anti phospho ikkα β ser176 180 16a6
Manufactured by Cell Signaling Technology
Sourced in United Kingdom, United States
Anti-phospho-IKKα/β (Ser176/180) (16A6) is a mouse monoclonal antibody that recognizes the phosphorylated forms of IKKα and IKKβ at Ser176 and Ser180 residues. This antibody is intended for use in western blotting applications.
Automatically generated - may contain errors
Lab products found in correlation
Anti phospho nf κb p65 ser536 93h1, by Cell Signaling Technology (2 mentions)
Myd88 d80f5, by Cell Signaling Technology (1 mentions)
Anti iκbα 44d4, by Cell Signaling Technology (1 mentions)
Anti phospho iκbα ser32 36 5a5, by Cell Signaling Technology (1 mentions)
Anti nf κb p65 d14e12, by Cell Signaling Technology (1 mentions)
Anti ikkα, by Cell Signaling Technology (1 mentions)
Anti ikkβ, by Cell Signaling Technology (1 mentions)
Anti phospho creb ser133, by Cell Signaling Technology (1 mentions)
Anti caspase 1, by Cell Signaling Technology (1 mentions)
Anti iκbα l35a5, by Cell Signaling Technology (1 mentions)
Anti c ebpβ, by Cell Signaling Technology (1 mentions)
Anti myd88 d80f5, by Cell Signaling Technology (1 mentions)
Anti traf6 d21g3, by Cell Signaling Technology (1 mentions)
Anti phospho c ebpβ thr235, by Cell Signaling Technology (1 mentions)
Enhanced chemi luminescence (ecl), by Thermo Fisher Scientific (1 mentions)
Peroxidase conjugated secondary antibodies, by BioLegend (1 mentions)
Anti nf κb p65, by Cell Signaling Technology (1 mentions)
Anti β actin, by Merck Group (1 mentions)
Anti iκbα, by BD (1 mentions)
Anti sharpin 14626 1 ap, by Proteintech (1 mentions)
4 protocols using anti phospho ikkα β ser176 180 16a6
1
Western Blotting of NF-κB Pathway
2
Analyzing Inflammasome Signaling in Neutrophils
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A total of 106 neutrophils were lysed and analyzed by Western blotting as previously described (26 (link)) using the following antibodies: anti-IL-1β (3ZD; National Cancer Institute Biological Resources), anti-NLRP3 (D2P5E; Cell Signaling), anti-caspase-1 (Cell Signaling), anti-NF-κB p65 (D14E12; Cell Signaling), anti-phospho-NF-κB p65 (Ser536) (93H1; Cell Signaling), anti-IκBα (L35A5; Cell Signaling), anti-MyD88 (D80F5; Cell Signaling), anti-TRAF6 (D21G3; Cell Signaling), anti-IKKα (Cell Signaling), anti-IKKβ (2C8; Cell Signaling), anti-phospho-IKKα/β (Ser176/180) (16A6; Cell Signaling), anti-CREB1 (48H2; Cell Signaling), anti-phospho-CREB (Ser133) (87G3; Cell Signaling), anti-C/EBPβ (Cell Signaling), anti-phospho-C/EBPβ (Thr235) (Cell Signaling), or anti-β-actin (AC-15; Sigma-Aldrich). Peroxidase-conjugated secondary antibodies were used (BioLegend). Membranes were developed using ECL (Thermo Scientific) and detected using a Nikon camera as previously described (79 (link)). Quantification analysis of blots was performed using ImageJ, and β-actin was used as a loading control. The results for samples were expressed as a percentage of the value for the positive control (LPS or LPS+ATP group).
3
Antibody Immunoblot Detection Protocol
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The antibodies used for detection on immunoblots were anti-HOIL-1 N-ter, provided by H. Walczak (University College London, London, England, UK; Haas et al., 2009 (link)), anti-HOIL-1 C-ter provided by K. Iwai (Graduate School of Medicine, Kyoto University, Kyoto, Japan; Kirisako et al., 2006 (link)), anti-HOIP (PAB6229; Abnova), and anti-linear polyubiquitin chains 1F113F5/Y102L provided by Genentech (Matsumoto et al., 2012 (link)), anti-NEMO (sc-8330; Santa Cruz Biotechnology, Inc.; #611306; BD), anti-SHARPIN (14626–1-AP; ProteinTech), anti-IκBα (#610690; BD), anti-phospho-IKKα-β (ser176/180; 16A6; Cell Signaling Technology), anti-IKKβ (AM8109a, Abgent), anti-β-tubulin (T4026; Sigma-Aldrich), and anti-GAPDH (sc-365062; Santa Cruz Biotechnology, Inc.) antibodies. Species-specific secondary antibodies coupled to horseradish peroxidase were obtained from Vector Laboratories or Amersham-Pharmacia.
4
Immunoblot Analysis of Inflammatory Signaling Pathways
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Anti-IκBα (C-21, 1:1000), anti-JNK2 (C-17,1:1000), and anti-IKKα (H-744, 1:1000) were from Santa Cruz Biotechnology (Dallas, TX, USA). Anti-phospho-IκBα (Ser32, 14D4, 1:1000), anti-phospho-p38 (Thr180/Tyr182, 3D7, 1:1000), phospho-JNK (Thr180/Tyr185, 1: 1000), anti-phospho-IKKα/β (Ser176/180, 16A6, 1:1000), and anti-ABIN-1 (1:1000) were purchased from Cell Signaling Technology (Danvers, MA, USA). Anti-actin (C-4, 1:10000) and lipopolysaccharides (LPS, Escherichia coli 055: B5) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Tumor necrosis factor (TNF) was purchased from Peptech (Burlington, MA, USA).
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