Recombinant Anti-BubR1 antibody [EPR20652] (ab209998)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR20652] to BubR1
- Suitable for: WB, ICC/IF, IP
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-BubR1 antibody [EPR20652]
See all BubR1 primary antibodies -
Description
Rabbit monoclonal [EPR20652] to BubR1 -
Host species
Rabbit -
Tested applications
Suitable for: WB, ICC/IF, IPmore 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: Human cell lines HeLa, Raji and HepG2 whole cell lysates. ICC/IF: Human cell lines HeLa, HepG2, A549 and HT-29. IP: Human cell line HeLa 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.
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 long term. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR20652 -
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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Recombinant Protein
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab209998 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 |
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WB |
1/1000. Detects a band of approximately 120 kDa (predicted molecular weight: 119 kDa).
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ICC/IF |
1/500.
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IP |
1/30.
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Notes |
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WB
1/1000. Detects a band of approximately 120 kDa (predicted molecular weight: 119 kDa). |
ICC/IF
1/500. |
IP
1/30. |
Target
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Function
Essential component of the mitotic checkpoint. Required for normal mitosis progression. The mitotic checkpoint delays anaphase until all chromosomes are properly attached to the mitotic spindle. One of its checkpoint functions is to inhibit the activity of the anaphase-promoting complex/cyclosome (APC/C) by blocking the binding of CDC20 to APC/C, independently of its kinase activity. The other is to monitor kinetochore activities that depend on the kinetochore motor CENPE. Required for kinetochore localization of CENPE. Negatively regulates PLK1 activity in interphase cells and suppresses centrosome amplification. Also implicated in triggering apoptosis in polyploid cells that exit aberrantly from mitotic arrest. May play a role for tumor suppression. -
Tissue specificity
Highly expressed in thymus followed by spleen. Preferentially expressed in tissues with a high mitotic index. -
Involvement in disease
Note=Defects in BUB1B are associated with tumor formation.
Defects in BUB1B are the cause of premature chromatid separation trait (PCS) [MIM:176430]. PCS consists of separate and splayed chromatids with discernible centromeres and involves all or most chromosomes of a metaphase. It is found in up to 2% of metaphases in cultured lymphocytes from approximately 40% of normal individuals. When PCS is present in 5% or more of cells, it is known as the heterozygous PCS trait and has no obvious phenotypic effect, although some have reported decreased fertility. Inheritance is autosomal dominant.
Defects in BUB1B are the cause of mosaic variegated aneuploidy syndrome (MVA) [MIM:257300]. MVA is a severe autosomal recessive developmental disorder characterized by mosaic aneuploidies, predominantly trisomies and monosomies, involving multiple different chromosomes and tissues. The proportion of aneuploid cells varies but is usually more than 25% and is substantially greater than in normal individuals. Affected individuals typically present with severe intrauterine growth retardation and microcephaly. Eye anomalies, mild dysmorphism, variable developmental delay, and a broad spectrum of additional congenital abnormalities and medical conditions may also occur. The risk of malignancy is high, with rhabdomyosarcoma, Wilms tumor and leukemia reported in several cases. MVA is caused by biallelic mutations in the BUB1B gene. -
Sequence similarities
Belongs to the protein kinase superfamily. Ser/Thr protein kinase family. BUB1 subfamily.
Contains 1 BUB1 N-terminal domain.
Contains 1 protein kinase domain. -
Domain
The D-box targets the protein for rapid degradation by ubiquitin-dependent proteolysis during the transition from mitosis to interphase.
The BUB1 N-terminal domain directs kinetochore localization and binding to BUB3. -
Post-translational
modificationsProteolytically cleaved by caspase-3 in a cell cycle specific manner. The cleavage might be involved in the durability of the cell cycle delay. Caspase-3 cleavage is associated with abrogation of the mitotic checkpoint. The major site of cleavage is at Asp-610.
Acetylation at Lys-250 regulates its degradation and timing in anaphase entry.
Ubiquitinated. Degradated by the proteasome.
Sumoylated by SUMO2 and SUMO3. The sumoylation mediates the association with CENPE at the kinetochore.
Autophosphorylated in vitro. Intramolecular autophosphorylation is stimulated by CENPE. Phosphorylated during mitosis and hyperphosphorylated in mitotically arrested cells. Phosphorylation at Ser-670 and Ser-1043 occurs at kinetochores upon mitotic entry with dephosphorylation at the onset of anaphase. -
Cellular localization
Cytoplasm. Nucleus. Chromosome > centromere > kinetochore. Cytoplasm > cytoskeleton > centrosome. Cytoplasmic in interphase cells. Associates with the kinetochores in early prophase. Kinetochore localization requires BUB1, PLK1 and CASC5. - Information by UniProt
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Database links
- Entrez Gene: 701 Human
- Omim: 602860 Human
- SwissProt: O60566 Human
- Unigene: 513645 Human
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Alternative names
- Beta homolg of S. cerevisiae BUB 1 antibody
- Beta homolg of S. cerevisiae budding uninhibited by benzimidazoles antibody
- BUB 1B antibody
see all
Images
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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HT-29 (human colorectal adenocarcinoma cell line) cells labeling BubR1 with ab209998 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing specific staining on centromeres of HT-29 cell line.
The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.
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BubR1 was immunoprecipitated from 0.35 mg of HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab209998 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab209998 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1/1000 dilution.
Lane 1: HeLa whole cell lysate 10 µg (Input).Lane 2: ab209998 IP in HeLa whole cell lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab209998 in HeLa whole cell lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 1 second.
All lanes : Anti-BubR1 antibody [EPR20652] (ab209998) at 1/1000 dilution
All lanes :
Secondary
All lanes : VeriBlot for IP Detection Reagent (HRP) (ab131366)
Observed band size: 120 kDa why is the actual band size different from the predicted?
Exposure time: 1 second -
Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A549 (human lung carcinoma cell line) cells labeling BubR1 with ab209998 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing specific staining on centromeres of A549 cell line.
The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.
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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HepG2 (human liver hepatocellular carcinoma cell line) cells labeling BubR1 with ab209998 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing specific staining on centromeres of HepG2 cell line.
The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.
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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling BubR1 with ab209998 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing subcellular localization of BubR1 changes through mitosis of HeLa cell line (PMID: 25833949). The nuclear counter stain is DAPI (blue).
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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling BubR1 with ab209998 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing specific staining on centromeres of HeLa cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] Microtubule Marker (Alexa Fluor® 594) (ab195889) at 1/200 dilution.
-ve control: Secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary at 1/1000 dilution. -
All lanes : Anti-BubR1 antibody [EPR20652] (ab209998) at 1/1000 dilution
Lane 1 : HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : Raji (human Burkitt's lymphoma cell line) whole cell lysate
Lane 3 : HepG2 (human liver hepatocellular carcinoma cell line) whole cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Performed under reducing conditions.
Predicted band size: 119 kDa
Observed band size: 120 kDa why is the actual band size different from the predicted?Positive BubR1 detection by WB is dependent on the number of mitotic cells in the sample and may need optimization for some cell and tissue lysates.
Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: Lane 1 - 10 seconds, lane 2 - 3 seconds, lane 3 - 3 minutes
Protocols
Datasheets and documents
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SDS download
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Datasheet download
Certificate of Compliance
References (0)
ab209998 has not yet been referenced specifically in any publications.