Immunohistochemical analysis of paraffin-embedded human breast carcinoma, using Phospho-NF-kappaB p105 (Ser932) (178F3) Rabbit mAb (IHC Specific) in the presence of control peptide (left) or Phospho-NF-kappaB p105 (Ser933) Blocking Peptide #1021 (right).
Immunohistochemical analysis of paraffin-embedded human colon carcinoma, showing cytoplasmic localization, using Phospho-NF-kappaB p105 (Ser932) (178F3) Rabbit mAb (IHC Specific).
Immunohistochemical analysis of paraffin-embedded human colon carcinoma untreated (left) or lambda-phosphatase-treated (right), using Phospho-NF-kappaB p105 (Ser932) (178F3) Rabbit mAb (IHC Specific).
Immunohistochemical analysis of paraffin-embedded human lung carcinoma, using Phospho-NF-kappaB p105 (Ser932) (178F3) Rabbit mAb (IHC Specific).
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
NOTE: Do not allow slides to dry at any time during this procedure.
For Citrate: Heat slides in a microwave submersed in 1X citrate unmasking solution until boiling is initiated; follow with 10 min at a sub-boiling temperature (95°-98°C). Cool slides on bench top for 30 min.
|SignalStain® Boost IHC Detection Reagent (HRP, Rabbit) #8114||SignalStain® Boost IHC Detection Reagent (AP, Rabbit) #18653|
|SignalStain® DAB Substrate Kit #8059||SignalStain® Vibrant Red Alkaline Phosphatase Substrate Kit #76713|
|SignalStain® Vivid Purple Peroxidase Substrate Kit #96632|
NOTE: Use of detection reagents other than those specified in this protocol may require further optimization of the primary antibody to account for the different sensitivities of the detection reagents.
posted June 2005
revised June 2020
Protocol Id: 310
Phospho-NF-kappaB p105 (Ser932) (178F3) Rabbit mAb detects endogenous levels of p105NF-kappaB only when phosphorylated at serine 932.
Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to amino acids around Ser932 of NF-kappaB p105.
Transcription factors of the nuclear factor κB (NF-κB)/Rel family play a pivotal role in inflammatory and immune responses (1,2). There are five family members in mammals: RelA, c-Rel, RelB, NF-κB1 (p105/p50), and NF-κB2 (p100/p52). Both p105 and p100 are proteolytically processed by the proteasome to produce p50 and p52, respectively. Rel proteins bind p50 and p52 to form dimeric complexes that bind DNA and regulate transcription. In unstimulated cells, NF-κB is sequestered in the cytoplasm by IκB inhibitory proteins (3-5). NF-κB-activating agents can induce the phosphorylation of IκB proteins, targeting them for rapid degradation through the ubiquitin-proteasome pathway and releasing NF-κB to enter the nucleus where it regulates gene expression (6-8). NIK and IKKα (IKK1) regulate the phosphorylation and processing of NF-κB2 (p100) to produce p52, which translocates to the nucleus (9-11).
Following IKK-mediated phosphorylation of p105 NF-kappaB at multiple sites (Ser921, 923, 927 and 932) on its carboxy-terminus, SCFbeta-TrCP mediated processing produces the 50 kDa active form p50 (12,13).
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