Recombinant
RabMAb

Recombinant Anti-Bmi1 (phospho T275) antibody [EPR19848] - BSA and Azide free (ab228458)

Overview

  • Product name

    Anti-Bmi1 (phospho T275) antibody [EPR19848] - BSA and Azide free
    See all Bmi1 primary antibodies
  • Description

    Rabbit monoclonal [EPR19848] to Bmi1 (phospho T275) - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, Dot blot, ICC/IF, Flow Cyt, IPmore details
  • Species reactivity

    Reacts with: Mouse, Human
  • Immunogen

    Synthetic peptide within Mouse Bmi1 aa 250 to the C-terminus (phospho Y275). The exact sequence is proprietary.
    Database link: P25916

  • Positive control

    • ICC/IF: HEK-293T cells transfected with DDDDK and mCherry-tagged mouse Bmi1 (exposed to UV light).
  • General notes

    Ab228458 is the carrier-free version of ab213723. 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.

    ab228458 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.??

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab228458 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 Use at an assay dependent concentration. Predicted molecular weight: 37 kDa.
Dot blot Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.
IP Use at an assay dependent concentration.

Target

  • Function

    Component of the Polycomb group (PcG) multiprotein PRC1 complex, a complex required to maintain the transcriptionally repressive state of many genes, including Hox genes, throughout development. PcG PRC1 complex acts via chromatin remodeling and modification of histones; it mediates monoubiquitination of histone H2A 'Lys-119', rendering chromatin heritably changed in its expressibility. In the PRC1 complex, it is required to stimulate the E3 ubiquitin-protein ligase activity of RNF2/RING2.
  • Sequence similarities

    Contains 1 RING-type zinc finger.
  • Post-translational
    modifications

    Monoubiquitinated (By similarity). May be polyubiquitinated; which does not lead to proteasomal degradation.
  • Cellular localization

    Nucleus. Cytoplasm.
  • Information by UniProt
  • Database links

  • Alternative names

    • B lymphoma Mo MLV insertion region (mouse) antibody
    • B lymphoma Mo MLV insertion region 1 homolog antibody
    • Bmi 1 antibody
    • BMI1 antibody
    • BMI1 polycomb ring finger oncogene antibody
    • BMI1_HUMAN antibody
    • Flvi 2/bmi 1 antibody
    • FLVI2/BMI1 antibody
    • MGC12685 antibody
    • Murine leukemia viral (bmi 1) oncogene homolog antibody
    • Oncogene BMI 1 antibody
    • PCGF 4 antibody
    • PCGF4 antibody
    • Polycomb complex protein BMI 1 antibody
    • Polycomb complex protein BMI-1 antibody
    • Polycomb group protein Bmi1 antibody
    • Polycomb group ring finger 4 antibody
    • Polycomb group RING finger protein 4 antibody
    • RING finger protein 51 antibody
    • RNF 51 antibody
    • RNF51 antibody
    see all

Images

  • Bmi1 (phospho T275) was immunoprecipitated from 0.35 mg of HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) (transfected with DDDDK and mCherry-tagged mouse Bmi1 (WT) expression vector, followed by treatment with 800 J/m² UV, then cultured in DMEM containing 10% FBS for 30 minutes) whole cell lysate with ab213723 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab213723 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

    Lane 1: HEK-293T (transfected / UV treated) whole cell lysate 10 μg (Input).

    Lane 2: ab213723 IP in HEK-293T (transfected / UV treated) lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab213723 in HEK-293T (transfected / UV treated) whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 10 seconds.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab213723).

  • Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) cell line transfected with DDDDK and mCherry-tagged mouse Bmi1 T275A mutant expression vector (left), or DDDDK and mCherry-tagged mouse Bmi1 WT expression vector (right) labeling Bmi1 (phospho T275) with ab213723 at 1/500 (red) compared with a Rabbit monoclonal IgG (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (black). 

    Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077), at 1/2000 dilution was used as the secondary antibody.

    The cells were gated on the mCherry positive population.

    The plasmids were kindly provided by Dr. Wei Guo from Tsinghua University.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab213723).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized U-2 OS (human bone osteosarcoma epithelial cell line) cells labeling Bmi1 (phospho T275) with ab213723 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing increased nuclear foci staining observed in UV-treated U-2 OS cells. The cells were treated with 50 J/m² UV, then cultured in McCoy's 5a media supplemented with 10% FBS for 2 hours.

    The nuclear counterstain is DAPI (blue). Tubulin is detected with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)  at 1/200 dilution (red).

    The negative controls are as follows:
    -ve control 1: PBS instead of primary antibody (non-treated cells), followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.
    -ve control 2: PBS instead of primary antibody (UV treated cells), followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

     

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab213723).

  • Dot blot analysis of Bmi1 (phospho T275) labeled with ab213723 at 1/1,000 dilution.

    Lane 1: Bmi1 (phospho T275) peptide.

    Lane 2: Bmi1 non-phospho peptide.

    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100,000 dilution was used as secondary antibody.

    Blocking and dilution buffer: 5% NFDM/TBST.

    Exposure time: 3 minutes.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab213723).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HEK-293T cells (human epithelial cell line from embryonic kidney transformed with large T antigen) transfected with DDDDK and mCherry-tagged mouse Bmi1 (WT) expression vector labeling Bmi1 (phospho T275) with ab213723 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining in HEK-293T cells transfected with DDDDK and mCherry-tagged mouse Bmi1 (WT) expression vector. No staining was observed in HEK-293T cells transfected with DDDDK and mCherry-tagged mouse Bmi1 T275A mutant expression vector.

    The nuclear counterstain is DAPI (blue). Tubulin is detected with Anti-Tubulin antibody [YOL1/34] - Microtubule Marker (Alexa Fluor® 647) (ab195884) at 1/200 dilution (white).

    The negative controls are as follows:
    -ve control 1: PBS instead of primary antibody (HEK-293T transfected with wild-type BMi1), followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.
    -ve control 2: PBS instead of primary antibody (HEK-293T transfected with Bmi1 T275A construct), followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

    Plasmids were kindly provided by Dr. Wei Guo from Tsinghua University.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab213723).

References

ab228458 has not yet been referenced specifically in any publications.

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