Recombinant Anti-IKK beta antibody [Y466] (ab32135)
![Western blot - Anti-IKK beta antibody [Y466] (ab32135)](https://www.abcam.com/ps/products/32/ab32135/Images/ab32135-449142-anti-ikk-beta-antibody-y466-western-blot-hela-edited-lysates.jpg "Western blot - Anti-IKK beta antibody [Y466] (ab32135)")
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [Y466] to IKK beta
- Suitable for: WB, IP
- Knockout validated
- Reacts with: Mouse, Human
Related conjugates and formulations
Overview
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Product name
Anti-IKK beta antibody [Y466]
See all IKK beta primary antibodies -
Description
Rabbit monoclonal [Y466] to IKK beta -
Host species
Rabbit -
Tested applications
Suitable for: WB, IPmore details -
Species reactivity
Reacts with: Mouse, Human -
Immunogen
Synthetic peptide within Human IKK beta aa 650-750 (C terminal). The exact sequence is proprietary.
Database link: O14920 -
Positive control
- WB: Wild-type HAP1 cell lysate. HeLa, NIH/3T3 and Daudi cell lysate. IHC-P: Human cervical carcinoma tissue. IP: SH-SY5Y whole cell lysate.
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General notes
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.
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.
Rat: We have preliminary internal testing data to indicate this antibody may not react with this species. Please contact us for more information.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol, 0.05% BSA -
Concentration information loading... -
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
Y466 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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KO cell lines
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KO cell lysates
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Positive Controls
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Recombinant Protein
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab32135 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Abreviews | Notes |
|---|---|---|
| WB | (3) |
1/1000 - 1/5000. Detects a band of approximately 87 kDa.
This antibody has weaker binding with mouse than human. |
| IP |
1/50.
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| Notes |
|---|
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WB
1/1000 - 1/5000. Detects a band of approximately 87 kDa. This antibody has weaker binding with mouse than human. |
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IP
1/50. |
Target
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Function
Acts 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. Also phosphorylates NCOA3. -
Tissue specificity
Highly expressed in heart, placenta, skeletal muscle, kidney, pancreas, spleen, thymus, prostate, testis and peripheral blood. -
Sequence similarities
Belongs to the protein kinase superfamily. Ser/Thr protein kinase family. I-kappa-B kinase subfamily.
Contains 1 protein kinase domain. -
Post-translational
modificationsUpon cytokine stimulation, phosphorylated on Ser-177 and Ser-181 by MEKK1 and/or MAP3K14/NIK; which enhances activity. Once activated, autophosphorylates on the C-terminal serine cluster; which decreases activity and prevents prolonged activation of the inflammatory response.
Acetylation of Thr-180 by Yersinia yopJ prevents phosphorylation and activation, thus blocking the I-kappa-B pathway.
Ubiquitinated. Monoubiquitination involves TRIM21 that leads to inhibition of Tax-induced NF-kappa-B signaling. According to PubMed:19675099, 'Ser-163' does not serve as a monoubiquitination site. According to PubMed:16267042, ubiquitination on 'Ser-163' modulates phosphorylation on C-terminal serine residues. Monoubiquitination by TRIM21 is dirupted by Yersinia yopJ. -
Cellular localization
Cytoplasm. Membrane raft. Colocalized with DPP4 in membrane rafts. - Information by UniProt
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Database links
- Entrez Gene: 3551 Human
- Entrez Gene: 16150 Mouse
- Omim: 603258 Human
- SwissProt: O14920 Human
- SwissProt: O88351 Mouse
- Unigene: 597664 Human
- Unigene: 277886 Mouse
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Alternative names
- I kappa B kinase 2 antibody
- I kappa B kinase beta antibody
- I-kappa-B kinase 2 antibody
see all
Images
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Western blot - Anti-IKK beta antibody [Y466] (ab32135)All lanes : Anti-IKK beta antibody [Y466] (ab32135) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : IKBKB CRISPR/Cas9 edited HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Observed band size: 87 kDa why is the actual band size different from the predicted?Lanes 1- 2: Merged signal (red and green). Green - ab32135 observed at 87 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab32135 was shown to react with IKK beta in wild-type HeLa cells in western blot. The band observed in CRISPR/Cas9 edited cell line ab264847 (CRISPR/Cas9 edited cell lysate ab257228) lane below 87kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Wild-type HeLa and IKBKB CRISPR/Cas9 edited HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab32135 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.
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![Immunoprecipitation - Anti-IKK beta antibody [Y466] (ab32135)](https://www.abcam.com/ps/products/32/ab32135/Images/ab32135-298014-anti-ikk-beta-antibody-y466-immunoprecipitation.jpg)
Immunoprecipitation - Anti-IKK beta antibody [Y466] (ab32135)ab32135 (purified) at 1:50 dilution (2μg) immunoprecipitating IKK beta in SH-SY5Y (Human neuroblastoma epithelial cell) whole cell lysate.
Lane 1 (input): SH-SY5Y (Human neuroblastoma epithelial cell) whole cell lysate 10ug
Lane 2 (+): ab32135 & SH-SY5Y (Human neuroblastoma epithelial cell) whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32135 in SH-SY5Y (Human neuroblastoma epithelial cell) whole cell lysateFor western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1:10,000 dilution.
Blocking and diluting buffer: 5% NFDM/TBST. -
![Western blot - Anti-IKK beta antibody [Y466] (ab32135)](https://www.abcam.com/ps/products/32/ab32135/Images/ab32135-252439-anti-ikk-beta-antibody-y466-western-blot.jpg)
Western blot - Anti-IKK beta antibody [Y466] (ab32135)Lanes 1, 3 and 5: Wild-type HAP1 cell lysate (20 µg)
Lanes 2, 4 and 6: IKK beta knockout HAP1 cell lysate (20 µg)
Lanes 1 and 2: Green signal from target – ab32135 observed at 87 kDa
Lanes 3 and 4: Red signal from loading control – ab8226 observed at 42 kDa
Lanes 5 and 6: Merged (red and green) signalUnpurified ab32135 was shown to react with IKK beta when IKK beta knockout samples were used, along with additional cross-reactive bands. Wild-type and IKK beta knockout samples were subjected to SDS-PAGE. ab32135 and ab8226 (loading control to beta actin) were diluted 1/1000 and 1/2000 respectively and incubated overnight at 4ºC. 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.
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![Western blot - Anti-IKK beta antibody [Y466] (ab32135)](https://www.abcam.com/ps/products/32/ab32135/Images/ab32135-296724-anti-ikk-beta-antibody-y466-western-blot.jpg)
Western blot - Anti-IKK beta antibody [Y466] (ab32135)All lanes : Anti-IKK beta antibody [Y466] (ab32135) at 1/1000 dilution (purified)
Lane 1 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysates
Lane 2 : Mouse lung lysates
Lane 3 : Rat brain lysates
Lane 4 : SH-SY5Y (Human neuroblastoma epithelial cell) whole cell lysates
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Observed band size: 87 kDa why is the actual band size different from the predicted?Blocking and diluting buffer: 5% NFDM/TBST
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![Western blot - Anti-IKK beta antibody [Y466] (ab32135)](https://www.abcam.com/ps/products/32/ab32135/Images/ab32135-296725-anti-ikk-beta-antibody-y466-western-blot.jpg)
Western blot - Anti-IKK beta antibody [Y466] (ab32135)Anti-IKK beta antibody [Y466] (ab32135) at 1/2500 dilution (purified) + HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates at 20 µg
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilutionBlocking and diluting buffer: 5% NFDM/TBST
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![Western blot - Anti-IKK beta antibody [Y466] (ab32135)](https://www.abcam.com/ps/products/32/ab32135/Images/ab32135-43995-anti-ikk-beta-antibody-y466-western-blot.jpg)
Western blot - Anti-IKK beta antibody [Y466] (ab32135)Anti-IKK beta antibody [Y466] (ab32135) at 1/5000 dilution (unpurified) + Daudi cell lysate
Observed band size: 87 kDa why is the actual band size different from the predicted? -
![Western blot - Anti-IKK beta antibody [Y466] (ab32135)](https://www.abcam.com/ps/products/32/ab32135/Images/ab32135-43993-anti-ikk-beta-antibody-y466-western-blot.jpg)
Western blot - Anti-IKK beta antibody [Y466] (ab32135)unpurified ab32135
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![Anti-IKK beta antibody [Y466] (ab32135)](https://www.abcam.com/ps/products/32/ab32135/Images/ab32135-9-benefits-of-recombinant-antibodies.png)
Anti-IKK beta antibody [Y466] (ab32135)
Protocols
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.
Datasheets and documents
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SDS download
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Datasheet download
References (38)
ab32135 has been referenced in 38 publications.
- He C et al. Modulatory Potential of LncRNA Zfas1 for Inflammation and Neuronal Apoptosis in Temporal Lobe Epilepsy. Yonsei Med J 62:215-223 (2021). PubMed: 33635011
- Zhang R et al. Ginkgolide C attenuates lipopolysaccharide-induced acute lung injury by inhibiting inflammation via regulating the CD40/NF-?B signaling pathway. Int J Mol Med 47:N/A (2021). PubMed: 33649807
- Saha S et al. Long Noncoding RNA DRAIC Inhibits Prostate Cancer Progression by Interacting with IKK to Inhibit NF-?B Activation. Cancer Res 80:950-963 (2020). PubMed: 31900260
- Qin T et al. A Novel Homozygous Mutation Destabilizes IKKß and Leads to Human Combined Immunodeficiency. Front Immunol 11:517544 (2020). PubMed: 33658989
- Quinn JA et al. The relationship between members of the canonical NF-kB pathway, tumour microenvironment and cancer specific survival in colorectal cancer patients. Histol Histopathol 35:569-578 (2020). PubMed: 31592535