Recombinant Anti-IKK alpha antibody [Y463] - BSA and Azide free (ab169743)
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 - BSA and Azide free
- Suitable for: ICC, Flow Cyt, IHC-P, IP, WB
- Knockout validated
- Reacts with: Mouse, Rat, Human
Overview
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Product name
Anti-IKK alpha antibody [Y463] - BSA and Azide free
See all IKK alpha primary antibodies -
Description
Rabbit monoclonal [Y463] to IKK alpha - BSA and Azide free -
Host species
Rabbit -
Tested Applications & Species
Application Species Flow Cyt HumanIHC-P HumanIP Human -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: Daudi cell lysate. IP: HeLa cell lysates
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General notes
Ab169743 is the carrier-free version of ab32041. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.
Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
ab169743 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.
Maxpar® is a trademark of Fluidigm Canada Inc.
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.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
Storage buffer
Constituent: PBS -
Carrier free
Yes -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
Y463 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Conjugation kits
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Isotype control
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Recombinant Protein
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab169743 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Tested applications are guaranteed to work and covered by our Abpromise guarantee.
Predicted to work for this combination of applications and species but not guaranteed.
Does not work for this combination of applications and species.
Application | Species |
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Flow Cyt |
Human
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IHC-P |
Human
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IP |
Human
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Application | Abreviews | Notes |
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ICC |
Use at an assay dependent concentration.
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Flow Cyt |
Use at an assay dependent concentration.
ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
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IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
|
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IP |
Use at an assay dependent concentration.
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WB |
Use at an assay dependent concentration. Detects a band of approximately 88 kDa (predicted molecular weight: 85 kDa).
|
Notes |
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ICC
Use at an assay dependent concentration. |
Flow Cyt
Use at an assay dependent concentration. ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
IP
Use at an assay dependent concentration. |
WB
Use at an assay dependent concentration. Detects a band of approximately 88 kDa (predicted molecular weight: 85 kDa). |
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. 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 specificity
Widely expressed. -
Involvement in disease
Defects 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 similarities
Belongs to the protein kinase superfamily. Ser/Thr protein kinase family. I-kappa-B kinase subfamily.
Contains 1 protein kinase domain. -
Post-translational
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 localization
Cytoplasm. Nucleus. Shuttles between the cytoplasm and the nucleus. - Information by UniProt
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Database links
- Entrez Gene: 1147 Human
- Entrez Gene: 309361 Rat
- Omim: 600664 Human
- SwissProt: O15111 Human
- SwissProt: Q60680 Mouse
- Unigene: 198998 Human
- Unigene: 3996 Mouse
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Alternative names
- chuk antibody
- CHUK1 antibody
- Conserved Helix Loop Helix Ubiquitous Kinase antibody
see all
Images
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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
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32041). -
Clone Y463 (ab169743) has been successfully conjugated by Abcam. This image was generated using Anti-IKK alpha antibody [Y463] (Alexa Fluor® 488). Please refer to ab200412 for protocol details.
ab200412 staining IKKα in wild-type HAP1 cells (top panel) and IKKα knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), 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 ab200412 at 1/400 dilution and ab7291 at 1μg/ml dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in pseudo colour red). Nuclear DNA was labelled in blue with DAPI.
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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.This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32041).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IKK alpha antibody [Y463] - BSA and Azide free (ab169743)
Ab32041, at a dilution of 1/50, staining IKK alpha in paraffin embedded human stomach carcinoma by Immunohistochemisty.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32041).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
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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.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32041).
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Clone Y463 (ab169743) has been successfully conjugated by Abcam. This image was generated using Anti-IKK alpha antibody [Y463] (PE). Please refer to ab210716 for protocol details.
ab210716 staining IKK alpha in HeLa cells. The cells were fixed with 100% methanol (5min), 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 overnight at +4°C with ab210716 at 1/50 dilution (pseudocolored in green) and ab195884, Rat monoclonal to Tubulin (Alexa Fluor® 647), at 1/250 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
This product also gave a positive signal under the same testing conditions in HeLa cells fixed with 4% formaldehyde (10 min).
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Clone Y463 (ab169743) has been successfully conjugated by Abcam. This image was generated using Anti-IKK alpha antibody [Y463] (Alexa Fluor® 647). Please refer to ab200414 for protocol details.
ab200414 staining IKK alpha in HeLa cells. 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 overnight at +4°C with ab200414 at 1/100 dilution (shown in red) and ab195887, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 488), at 2µg/ml (shown in green). Nuclear DNA was labelled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
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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 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 IgG (monoclonal) (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32041).
Protocols
Datasheets and documents
References (2)
ab169743 has been referenced in 2 publications.
- Kiss L et al. A tri-ionic anchor mechanism drives Ube2N-specific recruitment and K63-chain ubiquitination in TRIM ligases. Nat Commun 10:4502 (2019). PubMed: 31582740
- Clift D et al. A Method for the Acute and Rapid Degradation of Endogenous Proteins. Cell 171:1692-1706.e18 (2017). PubMed: 29153837