Recombinant Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)
![Western blot - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)](https://www.abcam.com/ps/products/247/ab247904/Images/ab247904-483146-ab109545369002antikap1antibodyepr5249westernblota431mcf7human.jpg "Western blot - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)")
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR5249] to KAP1 - BSA and Azide free
- Suitable for: IP, ICC/IF, WB, IHC-P, Flow Cyt (Intra)
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-KAP1 antibody [EPR5249] - BSA and Azide free
See all KAP1 primary antibodies -
Description
Rabbit monoclonal [EPR5249] to KAP1 - BSA and Azide free -
Host species
Rabbit -
Specificity
Although the immunogen is from a phosphor-peptide, this antibody detects phospho and non-phospho KAP1. Based on a peptide blocking experiment it has been found that the signal generated after non-phospho peptide blocking became much weaker, thus indicating that this antibody shows cross-reactivity with the non-phospho KAP1 at high level.
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Tested applications
Suitable for: IP, ICC/IF, WB, IHC-P, Flow Cyt (Intra)more details -
Species reactivity
Reacts with: Human -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: A431 and MCF7 cell lysates; IHC-P: Human colon and kidney tissue; ICC/IF: Hela cells; IP: A431 cell lysate. FC (Intra): HeLa cells.
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General notes
ab247904 is the carrier-free version of ab109545.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
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.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. 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
pH: 7.2
Constituent: PBS -
Carrier free
Yes -
Concentration information loading... -
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR5249 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Conjugation kits
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Isotype control
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Positive Controls
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Recombinant Protein
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab247904 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 |
|---|---|---|
| IP |
Use at an assay dependent concentration.
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| ICC/IF |
Use at an assay dependent concentration.
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| WB |
Use at an assay dependent concentration. Predicted molecular weight: 89 kDa.
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| IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
For antigen retrieval, heat up to 98 degree C, below boiling, and then let cool for 10-20 minutes.
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| Flow Cyt (Intra) |
Use at an assay dependent concentration.
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| Notes |
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IP
Use at an assay dependent concentration. |
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ICC/IF
Use at an assay dependent concentration. |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 89 kDa. |
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IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol. For antigen retrieval, heat up to 98 degree C, below boiling, and then let cool for 10-20 minutes.
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Flow Cyt (Intra)
Use at an assay dependent concentration. |
Target
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Function
Nuclear corepressor for KRAB domain-containing zinc finger proteins (KRAB-ZFPs). Mediates gene silencing by recruiting CHD3, a subunit of the nucleosome remodeling and deacetylation (NuRD) complex, and SETDB1 (which specifically methylates histone H3 at 'Lys-9' (H3K9me)) to the promoter regions of KRAB target genes. Enhances transcriptional repression by coordinating the increase in H3K9me, the decrease in histone H3 'Lys-9 and 'Lys-14' acetylation (H3K9ac and H3K14ac, respectively) and the disposition of HP1 proteins to silence gene expression. Recruitment of SETDB1 induces heterochromatinization. May play a role as a coactivator for CEBPB and NR3C1 in the transcriptional activation of ORM1. Also corepressor for ERBB4. Inhibits E2F1 activity by stimulating E2F1-HDAC1 complex formation and inhibiting E2F1 acetylation. May serve as a partial backup to prevent E2F1-mediated apoptosis in the absence of RB1. Important regulator of CDKN1A/p21(CIP1). Has E3 SUMO-protein ligase activity toward itself via its PHD-type zinc finger. -
Tissue specificity
Expressed in all tissues tested including spleen, thymus, prostate, testis, ovary, small intestine, colon and peripheral blood leukocytes. -
Pathway
Protein modification; protein sumoylation. -
Sequence similarities
Belongs to the TRIM/RBCC family.
Contains 2 B box-type zinc fingers.
Contains 1 bromo domain.
Contains 1 PHD-type zinc finger.
Contains 1 RING-type zinc finger. -
Domain
The HP1 box is both necessary and sufficient for HP1 binding.
The PHD-type zinc finger enhances CEBPB transcriptional activity. The PHD-type zinc finger, the HP1 box and the bromo domain, function together to assemble the machinery required for repression of KRAB domain-containing proteins. Acts as an intramolecular SUMO E3 ligase for autosumoylation of bromodomain.
The RING-finger-B Box-coiled-coil/tripartite motif (RBCC/TRIM motif) is required for interaction with the KRAB domain of KRAB-zinc finger proteins. Binds four zinc ions per molecule. The RING finger and the N-terminal of the leucine zipper alpha helical coiled-coil region of RBCC are required for oligomerization.
Contains one Pro-Xaa-Val-Xaa-Leu (PxVxL) motif, which is required for interaction with chromoshadow domains. This motif requires additional residues -7, -6, +4 and +5 of the central Val which contact the chromoshadow domain. -
Post-translational
modificationsPhosphorylated upon DNA damage, probably by ATM or ATR. ATM-induced phosphorylation on Ser-824 represses sumoylation leading to the de-repression of expression of a subset of genes involved in cell cycle control and apoptosis in response to genotoxic stress. Dephosphorylation by the phosphatases, PPP1CA and PP1CB forms, allows sumoylation and expression of TRIM28 target genes.
Sumoylation/desumoylation events regulate TRIM28-mediated transcriptional repression. Sumoylation is required for interaction with CHD3 and SETDB1 and the corepressor activity. Represses and is repressed by Ser-824 phosphorylation. Enhances the TRIM28 corepressor activity, inhibiting transcriptional activity of a number of genes including GADD45A and CDKN1A/p21. Lys-554, Lys-779 and Lys-804 are the major sites of sumoylation. In response to Dox-induced DNA damage, enhanced phosphorylation on Ser-824 prevents sumoylation and allows de-repression of CDKN1A/p21. -
Cellular localization
Nucleus. Associated with centromeric heterochromatin during cell differentiation through CBX1. - Information by UniProt
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Database links
- Entrez Gene: 10155 Human
- Omim: 601742 Human
- SwissProt: Q13263 Human
- Unigene: 467408 Human
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Alternative names
- E3 SUMO protein ligase TRIM28 antibody
- E3 SUMO-protein ligase TRIM28 antibody
- FLJ29029 antibody
see all
Images
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Western blot - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)All lanes : Anti-KAP1 antibody [EPR5249] (ab109545) at 1/10000 dilution
Lane 1 : A431 (Human epidermoid carcinoma epithelial cell) whole cell lysates
Lane 2 : MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates
Lysates/proteins at 15 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Developed using the ECL technique.
Predicted band size: 89 kDa
Exposure time: 10 secondsThis data was developed using ab109545, the same antibody clone in a different buffer formulation:
Blocking/diluting buffer and concentration: 5% NFDM/TBST
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![Flow Cytometry (Intracellular) - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)](https://www.abcam.com/ps/products/247/ab247904/Images/ab247904-483145-ab109545122495antikap1antibodyepr5249flowcytometry.jpg)
Flow Cytometry (Intracellular) - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)This data was developed using ab109545, the same antibody clone in a different buffer formulation:
Overlay histogram showing HeLa cells stained with ab109545 (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 (ab190545, 1/100 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.
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![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)](https://www.abcam.com/ps/products/247/ab247904/Images/ab247904-483144-ab109545122497antikap1antibodyepr5249immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)This data was developed using ab109545, the same antibody clone in a different buffer formulation:
ab109545, at a 1/100 dilution, staining KAP1 in formalin-fixed, paraffin-embedded Human kidney tissue.
Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
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![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)](https://www.abcam.com/ps/products/247/ab247904/Images/ab247904-483143-ab109545122498antikap1antibodyepr5249immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)This data was developed using ab109545, the same antibody clone in a different buffer formulation:
ab109545, at a 1/100 dilution, staining KAP1 in formalin-fixed, paraffin-embedded Human colon tissue.
Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
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![Immunocytochemistry/ Immunofluorescence - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)](https://www.abcam.com/ps/products/247/ab247904/Images/ab247904-483141-ab109545122496antikap1antibodyepr5249immunofluorescence.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904) -
![Western blot - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)](https://www.abcam.com/ps/products/247/ab247904/Images/ab247904-483048-ab109545290396antikap1antibodyepr5249westernblot.jpg)
Western blot - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)This data was developed using ab109545, the same antibody clone in a different buffer formulation:
Lane 1: Wild type HAP1 whole cell lysate (20 µg)
Lane 2: HAP1 + DMSO whole cell lysate (20 µg)
Lane 3: HAP1 + Blaomycin whole cell lysate (20 µg)
Lane 4: TRIM28 knockout HAP1 whole cell lysate (20 µg)
Lane 5: TRIM28 knockout HAP1 + DMSO whole cell lysate (20 µg)
Lane 6: TRIM28 knockout HAP1 + Blaomycin whole cell lysate (20 µg)
Lane 7: HeLa + DMSO whole cell lysate (20 µg)
Lane 8: HeLa + Blaomycin whole cell lysate (20 µg)Lanes 1 - 8: Merged signal (red and green). Green - ab109545 observed at 110 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab109545 was shown to specifically react with KAP1 in wild type cells as signal was lost in KAP1 knockout cells. Wild-type and KAP1 knockout samples were subjected to SDS-PAGE. ab109545 and ab8245 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 10000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
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![Immunoprecipitation - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)](https://www.abcam.com/ps/products/247/ab247904/Images/ab247904-1-anti-kap1-antibody-epr5249-immunoprecipitation-a4315.jpg)
Immunoprecipitation - Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)KAP1 was immunoprecipitated from 0.35 mg A431 (Human epidermoid carcinoma epithelial cell) cell lysate 10 µg with ab109545 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab109545 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: A431 (Human epidermoid carcinoma epithelial cell) cell lysate 10 µg
Lane 2: ab109545 IP in A431 cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab109545 in A431 cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 32 seconds
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109545).
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![Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)](https://www.abcam.com/ps/products/247/ab247904/Images/ab247904-2-benefits-of-recombinant-antibodies.png)
Anti-KAP1 antibody [EPR5249] - BSA and Azide free (ab247904)
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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Datasheet download
Certificate of Compliance
References (0)
ab247904 has not yet been referenced specifically in any publications.
