Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [Y463] to IKK alpha
- Suitable for: WB, IHC-P, Flow Cyt, IP, ICC/IF
- Knockout validated
- Reacts with: Mouse, Rat, Human
Product nameAnti-IKK alpha antibody [Y463]
See all IKK alpha primary antibodies
DescriptionRabbit monoclonal [Y463] to IKK alpha
Tested applicationsSuitable for: WB, IHC-P, Flow Cyt, IP, ICC/IFmore details
Species reactivityReacts with: Mouse, Rat, Human
Synthetic peptide within Human IKK alpha aa 1-100 (N terminal). The exact sequence is proprietary.
- WB: Daudi and wild-type HAP1 cell lysates. IHC-P: Human stomach carcinoma. ICC/IF: Wild-type HAP1 cells. Flow Cyt: HeLa cells.HAP1-WT cells. IP: HeLa cell lysate
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Storage instructionsShipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
Storage bufferpH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 49% PBS, 50% Glycerol, 0.05% BSA
Concentration information loading...
- Anti-IKK alpha antibody [Y463] - BSA and Azide free (ab169743)
- Anti-IKK alpha antibody [Y463] (Alexa Fluor® 488) (ab200412)
- Anti-IKK alpha antibody [Y463] (Alexa Fluor® 647) (ab200414)
- Anti-IKK alpha antibody [Y463] (HRP) (ab200415)
- Anti-IKK alpha antibody [Y463] (PE) (ab210716)
- Anti-IKK alpha antibody [Y463] (APC) (ab221910)
Our Abpromise guarantee covers the use of ab32041 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||1/10000 - 1/50000. Detects a band of approximately 88 kDa (predicted molecular weight: 85 kDa).|
|IHC-P||Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.|
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
FunctionActs as part of the IKK complex in the conventional pathway of NF-kappa-B activation and phosphorylates inhibitors of NF-kappa-B thus leading to the dissociation of the inhibitor/NF-kappa-B complex and ultimately the degradation of the inhibitor. As part of the non-canonical pathway of NF-kappa-B activation, the MAP3K14-activated CHUK/IKKA homodimer phosphorylates NFKB2/p100 associated with RelB, inducing its proteolytic processing to NFKB2/p52 and the formation of NF-kappa-B RelB-p52 complexes. Also phosphorylates NCOA3. Phosphorylates 'Ser-10' of histone H3 at NF-kappa-B-regulated promoters during inflammatory responses triggered by cytokines.
Tissue specificityWidely expressed.
Involvement in diseaseDefects in CHUK are the cause of cocoon syndrome (COCOS) [MIM:613630]; also known as fetal encasement syndrome. COCOS is a lethal syndrome characterized by multiple fetal malformations including defective face and seemingly absent limbs, which are bound to the trunk and encased under the skin.
Sequence similaritiesBelongs to the protein kinase superfamily. Ser/Thr protein kinase family. I-kappa-B kinase subfamily.
Contains 1 protein kinase domain.
modificationsPhosphorylated by MAP3K14/NIK, AKT and to a lesser extent by MEKK1, and dephosphorylated by PP2A. Autophosphorylated.
Acetylation of Thr-179 by Yersinia yopJ prevents phosphorylation and activation, thus blocking the I-kappa-B signaling pathway.
Cellular localizationCytoplasm. Nucleus. Shuttles between the cytoplasm and the nucleus.
- Information by UniProt
- chuk antibody
- CHUK1 antibody
- Conserved Helix Loop Helix Ubiquitous Kinase antibody
Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: IKK alpha knockout HAP1 cell lysate (20 µg)
Lanes 1 and 2: Merged signal (red and green). Green - ab32041 observed at 84 kDa. Red - loading control, ab8226, observed at 42 kDa.
ab32041 was shown to specifically react with IKK alpha when IKK alpha knockout samples were used. Wild-type and IKK alpha knockout samples were subjected to SDS-PAGE. ab32041 and ab8226 (loading control to beta actin) were diluted 1/10 000 and 1/1000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10 000 dilution for 1 h at room temperature before imaging.
Overlay histogram showing HAP1 wildtype (green line) and HAP1-CHUK knockout cells (red line) stained with ab32041. The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab32041, 1µg/ml) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) presorbed (ab150081) at 1/2000 dilution for 30 min at 22°C.
A rabbit IgG isotype control antibody (ab172730) was used at the same concentration and conditions as the primary antibody (HAP1 wildtype - black line, HAP1-CHUK knockout - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).
Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.
This antibody can also be used in HAP1 cells fixed with80% methanol (5 min), , permeabilized with 0.1% PBS-Triton X-100 for 15 min under the same conditions.
ab32041 staining IKK alpha in wild-type HAP1 cells (top panel) and IKK alpha knockout HAP1 cells (bottom panel). The cells were fixed with 4% formaldehyde (10min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab32041 at 1/500 dilution and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.
Purified ab32041 at 1/60 dilution (2µg) immunoprecipitating IKK alpha in HeLa whole cell lysate.
Lane 1 (input): HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate 10µg
Lane 2 (+): ab32041 + HeLa whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab169743 in HeLa whole cell lysate.
VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/1000 dilution) was used for Western blotting.
Blocking Buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM/TBST.
Observed band size: 88 kDa
Anti-IKK alpha antibody [Y463] (ab32041) at 1/50000 dilution + Daudi cell lysate
Predicted band size: 85 kDa
Observed band size: 88 kDa why is the actual band size different from the predicted?
Ab32041, at a dilution of 1/50, staining IKK alpha in paraffin embedded human stomach carcinoma by Immunohistochemisty.
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Overlay histogram showing HeLa cells stained with ab32041 (red line). The cells were fixed with 80% 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 (ab32041, 1/50 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
ab32041 has been referenced in 28 publications.
- Tang H et al. CDK5 inhibition in vitro and in vivo induces cell death in myeloma and overcomes the obstacle of bortezomib resistance. Int J Mol Med 45:1661-1672 (2020). PubMed: 32236619
- Ma Y et al. 1-Deoxynojirimycin in Mulberry (Morus indica L.) Leaves Ameliorates Stable Angina Pectoris in Patients With Coronary Heart Disease by Improving Antioxidant and Anti-inflammatory Capacities. Front Pharmacol 10:569 (2019). PubMed: 31164826
- Yang BY et al. Datura Metel L. Ameliorates Imiquimod-Induced Psoriasis-Like Dermatitis and Inhibits Inflammatory Cytokines Production through TLR7/8-MyD88-NF-?B-NLRP3 Inflammasome Pathway. Molecules 24:N/A (2019). PubMed: 31181689
- Colomer C et al. IKKa Kinase Regulates the DNA Damage Response and Drives Chemo-resistance in Cancer. Mol Cell 75:669-682.e5 (2019). PubMed: 31302002
- Chen Y et al. Epigenetically upregulated oncoprotein PLCE1 drives esophageal carcinoma angiogenesis and proliferation via activating the PI-PLCe-NF-?B signaling pathway and VEGF-C/ Bcl-2 expression. Mol Cancer 18:1 (2019). PubMed: 30609930