Anti-Bmi1 antibody [F6] - ChIP Grade (ab14389)

Knockout Tested Mouse monoclonal Bmi1 antibody [F6]. Validated in WB, IP, IHC, ICC, Flow Cyt, ChIP, ICC/IF and tested in Mouse, Rat, Rabbit, Cat, Human, Xenopus laevis, Zebrafish.

Overview

  • Product name
    Anti-Bmi1 antibody [F6] - ChIP Grade
    See all Bmi1 primary antibodies
  • Description
    Mouse monoclonal [F6] to Bmi1 - ChIP Grade
  • Host species
    Mouse
  • Specificity
    Recognizes mouse Bmi-1(triplet). The clone number has been updated from (1.T.21) to (F6) both clone numbers name the same antibody clone.
  • Tested applications
    Suitable for: ICC/IF, Flow Cyt, ChIP, WB, ICC, IP, IHC-Fr, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Rabbit, Cat, Human, Xenopus laevis, Zebrafish
  • Immunogen

    Recombinant fragment, corresponding to amino acids 1-202 of Mouse Bmi1

  • Positive control
    • U2OS whole cell lysate.

Properties

Applications

Our Abpromise guarantee covers the use of ab14389 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
ICC/IF Use a concentration of 0.65 µg/ml. PubMed: 19001505
Flow Cyt Use a concentration of 0.65 µg/ml. PubMed: 19001505

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

ChIP Use at an assay dependent concentration. PubMed: 18332116
WB Use a concentration of 0.2 - 2 µg/ml. Predicted molecular weight: 37 kDa. Detects Bmi-1 in RIPA lysates from U2OS cells. U2OS cell lysate was resolved by electrophoresis, transferred to nitrocellulose and probed with ab14389 0.2ug/ml. Proteins were visualized using a goat anti-mouse IgG labeled with HRP and a chemiluminescence detection system.
ICC Use at an assay dependent concentration.
IP Use at an assay dependent concentration.
IHC-Fr Use at an assay dependent concentration. PubMed: 17210912
IHC-P 1/100.

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

  • Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: Bmi1 knockout HAP1 cell lysate)
    Lane 3: U2OS cell lysate (40 ug)
    Lane 4: Molt-4 cell lysate (40ug)
    Lanes 1 - 4: Merged signal (red and green). Green - ab14389 observed at 42 kDa. Red - loading control, ab176560, observed at 52 kDa.

    ab14389 was shown to recognize Bmi1 when Bmi1 knockout samples were used, along with additional cross-reactive bands. Wild-type and Bmi1 knockout samples were subjected to SDS-PAGE. ab14389 at a concentration of 1 μg/ml and ab176560 (loading control to alpha tubulin) diluted to 1/10000 were incubated overnight at 4°C. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

  • SK-N-SH cells were fixed in 4% paraformaldehyde, permeabilized in 0.55 Triton-X100 and incubated for 1 hour with ab14389 (1/200). The Bmi1 staining is shown in red. The cells were counterstained with DAPI (blue). 100x magnification.

  • Cells were lysed in RIPA buffer. Samples (25 µg) of total protein were separated by SDS-polyacrylamide gel electrophoresis gel (PAGE) in a 12% gel and transferred onto a nitrocellulose membrane. This was followed by an incubation with mouse anti-BMI1 (ab14389) and a sheep anti-mouse HRP conjugated antibody diluted 1:2000 was used as a secondary antibody. HRP conjugated anti-GAPDH antibody (ab9482) was used as a loading control.

    The mCbx7 expressing cells were infected with lentivirus-based shRNA vectors targeting BMI1 or an irrelevant control (C).

  • Formalin-fixed paraffin-embedded (FFPE) colorectal cancer tissues and normal colorectal tissues. Endogenous peroxidase was blocked by incubating the sections in a 0.3% solution of hydrogen peroxide (in PBS) for 20 min. Antigen retrieval was performed by heating the sections for 10 min at 95°C in a citrate buffer. Tissues were incubated with ab14389 overnight (16 hrs). Staining was visualized using the Dako REAL EnVision Detection System, Peroxidase/DAB+, Rabbit/Mouse (Dako). The stained TMA sections were scanned using a 20x magnification on the semi-automated Ariol system (Leica Microsystems, Wetzlar, Germany). Tumor cell areas (tumor tissues) and colon epithelium (in normal tissues) were identified for positive (indicated by yellow dots) and negative (blue dots) nuclei in tumor cores. TMA slides were scanned using a 20x magnification. Shown are positively stained tumor cores (top image) and negative tumor cores (bottom image).

  • Western blot analysis of RIPA lysates from U2OS cells labelling Bmi-1 with ab14389 at 0.2μg . An IgG (HRP) goat anti-mouse was used as the secondary antibody.

     

  • ab14389 staining Bmi1 in human head and neck squamous cell cancers (HNSCC) by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections).
    Sections were deparaffinized in xylene, dehydrated through graded alcohols, and placed in 0.1% hydrogen peroxide to quench any endogenous peroxidase activity. A 5 minute, 750 W microwave pretreatment in citrate buffer (pH 6.0) was repeated 4 times and followed by treatment with 10% normal rabbit serum for 30 minutes to block nonspecific antibody binding. The slides were then incubated with ab14389 at a 1/100. Primary antibody incubation was followed by three washes with PBST and incubation with fluorescently-labeled Cy2 (1/250) secondary antibody for 2 hours. Nuclei were counterstained with DAPI. Finally, sections were rinsed with PBST, dehydrated through sequential washes in 50%, 70%, 95%, and 100% ethanol and then cleared in xylene. Slides were mounted with DPX mounting media and allowed to dry overnight.

References

This product has been referenced in:
  • Zheng Q  et al. IL-17A promotes cell migration and invasion of glioblastoma cells via activation of PI3K/AKT signalling pathway. J Cell Mol Med 23:357-369 (2019). Read more (PubMed: 30353649) »
  • Chow HY  et al. Group I Paks are essential for epithelial- mesenchymal transition in an Apc-driven model of colorectal cancer. Nat Commun 9:3473 (2018). Read more (PubMed: 30150766) »
See all 56 Publications for this product

Customer reviews and Q&As

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1-10 of 12 Abreviews

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (differentiated induced pluripotent stem cells)
Permeabilization
Yes - 0.25% Triton X-100 in PBS
Specification
differentiated induced pluripotent stem cells
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: 22°C
Fixative
Formaldehyde

Abcam user community

Verified customer

Submitted Jul 19 2017

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Rhesus monkey Tissue sections (Jejunum)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Ventana CC1 (High pH)
Permeabilization
No
Specification
Jejunum
Fixative
Paraformaldehyde

Dr. Li Li

Verified customer

Submitted May 31 2017

Application
Western blot
Sample
Human Cell lysate - whole cell (human acute promylocyitc leukemia NB4 cells)
Loading amount
40 µg
Specification
human acute promylocyitc leukemia NB4 cells
Treatment
1 uM ATO treatment for indicated time
Gel Running Conditions
Reduced Denaturing (10% gel)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Mr. Sungsin Jo

Verified customer

Submitted Apr 02 2013

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Brain)
Specification
Brain
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citrate buffer pH6
Permeabilization
No
Blocking step
Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 7.5% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Oct 02 2012

Application
Western blot
Sample
Mouse Tissue lysate - whole (pancreatic cancer sample)
Loading amount
50 µg
Specification
pancreatic cancer sample
Gel Running Conditions
Reduced Denaturing
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Mar 20 2012

Application
Western blot
Sample
Human Cell lysate - whole cell (Panc1 cancer cell)
Loading amount
35 µg
Specification
Panc1 cancer cell
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Mar 20 2012

Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (skin section)
Specification
skin section
Fixative
Acetone
Permeabilization
Yes - TBST
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Mar 20 2012

Application
Immunohistochemistry (Frozen sections)
Sample
Human Tissue sections (SSC sections)
Specification
SSC sections
Fixative
Acetone
Permeabilization
Yes - TBST
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Mar 20 2012

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Cat Tissue sections (skin, spleen, small intestine)
Specification
skin, spleen, small intestine
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: citrate buffer
Permeabilization
No
Blocking step
universal agent as blocking agent for 10 minute(s) · Concentration: 0% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Oct 13 2011

Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (putative stem cell isoleated from pancreatic tumor)
Specification
putative stem cell isoleated from pancreatic tumor
Fixative
Aceton:Methanol 1:1
Permeabilization
Yes - PBS + Triton 0.025%
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Oct 01 2010

1-10 of 12 Abreviews

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