Anti-NFkB p105 / p50 antibody [E381] (ab32360)
- Product nameAnti-NFkB p105 / p50 antibody [E381]See all NFkB p105 / p50 primary antibodies ...
- DescriptionRabbit monoclonal [E381] to NFkB p105 / p50
- SpecificityThis antibody will detect both forms: p50 and p105.
- Tested applicationsWB, IHC-P, ICC, Flow Cyt more details
- Species reactivityReacts with: Mouse, Rat, Human
A synthetic peptide corresponding to residues in the nuclear factor NFkB p50 subunit.
- Positive control
- HeLa cell lysate; human prostate carcinoma
- General notes
Produced under U.S. Patent No. 5,675,063.
- Storage instructionsShipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
- Storage bufferPBS 49%,Sodium azide 0.01%,Glycerol 50%,BSA 0.05%
- Concentration information loading...
- PurityProtein A purified
- Clonality Monoclonal
- Clone numberE381
Our Abpromise guarantee covers the use of ab32360 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||WB: 1/1000 - 1/10000. Detects a band of approximately 50, 105 kDa (predicted molecular weight: 50 kDa).|
|IHC-P||IHC-P: 1/250 - 1/500.|
|ICC||ICC: 1/250 - 1/500.|
|Flow Cyt||Flow Cyt: 1/30.|
- FunctionNF-kappa-B is a pleiotropic transcription factor which is present in almost all cell types and is involved in many biological processed such as inflammation, immunity, differentiation, cell growth, tumorigenesis and apoptosis. NF-kappa-B is a homo- or heterodimeric complex formed by the Rel-like domain-containing proteins RELA/p65, RELB, NFKB1/p105, NFKB1/p50, REL and NFKB2/p52 and the heterodimeric p65-p50 complex appears to be most abundant one. The dimers bind at kappa-B sites in the DNA of their target genes and the individual dimers have distinct preferences for different kappa-B sites that they can bind with distinguishable affinity and specificity. Different dimer combinations act as transcriptional activators or repressors, respectively. NF-kappa-B is controlled by various mechanisms of post-translational modification and subcellular compartmentalization as well as by interactions with other cofactors or corepressors. NF-kappa-B complexes are held in the cytoplasm in an inactive state complexed with members of the NF-kappa-B inhibitor (I-kappa-B) family. In a conventional activation pathway, I-kappa-B is phosphorylated by I-kappa-B kinases (IKKs) in response to different activators, subsequently degraded thus liberating the active NF-kappa-B complex which translocates to the nucleus. NF-kappa-B heterodimeric p65-p50 and RelB-p50 complexes are transcriptional activators. The NF-kappa-B p50-p50 homodimer is a transcriptional repressor, but can act as a transcriptional activator when associated with BCL3. NFKB1 appears to have dual functions such as cytoplasmic retention of attached NF-kappa-B proteins by p105 and generation of p50 by a cotranslational processing. The proteasome-mediated process ensures the production of both p50 and p105 and preserves their independent function, although processing of NFKB1/p105 also appears to occur post-translationally. p50 binds to the kappa-B consensus sequence 5'-GGRNNYYCC-3', located in the enhancer region of genes involved in immune response and acute phase reactions. In a complex with MAP3K8, NFKB1/p105 represses MAP3K8-induced MAPK signaling; active MAP3K8 is released by proteasome-dependent degradation of NFKB1/p105.
- Sequence similaritiesContains 7 ANK repeats.
Contains 1 death domain.
Contains 1 RHD (Rel-like) domain.
- DomainThe C-terminus of p105 might be involved in cytoplasmic retention, inhibition of DNA-binding, and transcription activation.
Glycine-rich region (GRR) appears to be a critical element in the generation of p50.
modificationsWhile translation occurs, the particular unfolded structure after the GRR repeat promotes the generation of p50 making it an acceptable substrate for the proteasome. This process is known as cotranslational processing. The processed form is active and the unprocessed form acts as an inhibitor (I kappa B-like), being able to form cytosolic complexes with NF-kappa B, trapping it in the cytoplasm. Complete folding of the region downstream of the GRR repeat precludes processing.
Phosphorylation at 'Ser-903' and 'Ser-907' primes p105 for proteolytic processing in response to TNF-alpha stimulation. Phosphorylation at 'Ser-927' and 'Ser-932' are required for BTRC/BTRCP-mediated proteolysis.
Polyubiquitination seems to allow p105 processing.
S-nitrosylation of Cys-61 affects DNA binding.
- Cellular localizationNucleus. Cytoplasm. Nuclear, but also found in the cytoplasm in an inactive form complexed to an inhibitor.
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Anti-NFkB p105 / p50 antibody [E381] images
Anti-NFkB p105 / p50 antibody [E381] (ab32360) at 1/5000 dilution + HeLa cell lysate
Predicted band size : 50 kDa
Observed band size : 50,105 kDa (why is the actual band size different from the predicted?)
Paraffin-embedded human prostate carcinoma
ab32360 at 1/250 dilution
ICC/IF image of ab32360 stained MCF7 cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab32360, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
Overlay histogram showing K562 cells stained with ab32360 (red line). The cells were fixed with methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab32360, 1/50 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit monoclonal IgG (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a decreased signal in K562 cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween used under the same conditions.
References for Anti-NFkB p105 / p50 antibody [E381] (ab32360)
This product has been referenced in:
- Tordella L et al. ASPP2 suppresses squamous cell carcinoma via RelA/p65-mediated repression of p63. Proc Natl Acad Sci U S A 110:17969-74 (2013). WB ; Mouse . Read more (PubMed: 24127607) »
- Hong EJ et al. Loss of estrogen-related receptor a promotes hepatocarcinogenesis development via metabolic and inflammatory disturbances. Proc Natl Acad Sci U S A 110:17975-80 (2013). WB ; Mouse . Read more (PubMed: 24127579) »