Overview

  • Product nameAnti-CENPF antibody
    See all CENPF primary antibodies
  • Description
    Rabbit polyclonal to CENPF
  • SpecificityThis antibody is specific for CENP-F.
  • Tested applicationsSuitable for: Flow Cyt, ICC/IF, ICC, IHC-P, WB, IPmore details
  • Species reactivity
    Reacts with: Mouse, Human
  • Immunogen

    Fusion protein with C-terminus of CENP-F (Human).

  • Positive control
    • WB: Hela whole cell lysate ICC: Hela cells, SK-N-SH cells.

Properties

Applications

Our Abpromise guarantee covers the use of ab5 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
Flow Cyt 1/200.
ICC/IF 1/400 - 1/750.
ICC Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration.
WB 1/1500. Detects a band of approximately 330 kDa (predicted molecular weight: 330 kDa). Block membranes for 1 hour with 5% nonfat dry milk/TBS-T.
IP 1/50.

Target

  • FunctionRequired for kinetochore function and chromosome segregation in mitosis. Required for kinetochore localization of dynein, LIS1, NDE1 and NDEL1. Regulates recycling of the plasma membrane by acting as a link between recycling vesicles and the microtubule network though its association with STX4 and SNAP25. Acts as a potential inhibitor of pocket protein-mediated cellular processes during development by regulating the activity of RB proteins during cell division and proliferation. May play a regulatory or permissive role in the normal embryonic cardiomyocyte cell cycle and in promoting continued mitosis in transformed, abnormally dividing neonatal cardiomyocytes. Interaction with RB directs embryonic stem cells toward a cardiac lineage. Involved in the regulation of DNA synthesis and hence cell cycle progression, via its C-terminus. Has a potential role regulating skeletal myogenesis and in cell differentiation in embryogenesis. Involved in dendritic cell regulation of T-cell immunity against chlamydia.
  • Involvement in diseaseStromme syndrome
  • Sequence similaritiesBelongs to the centromere protein F family.
  • Developmental stageGradually accumulates during the cell cycle, reaching peak levels in G2 and M phase, and is rapidly degraded upon completion of mitosis.
  • Post-translational
    modifications
    Hyperphosphorylated during mitosis.
  • Cellular localizationCytoplasm, perinuclear region. Nucleus matrix. Chromosome, centromere, kinetochore. Cytoplasm, cytoskeleton, spindle. Relocalizes to the kinetochore/centromere (coronal surface of the outer plate) and the spindle during mitosis. Observed in nucleus during interphase but not in the nucleolus. At metaphase becomes localized to areas including kinetochore and mitotic apparatus as well as cytoplasm. By telophase, is concentrated within the intracellular bridge at either side of the mid-body.
  • Information by UniProt
  • Database links
  • Alternative names
    • AH antigen antibody
    • Cell cycle dependent 350K nuclear protein antibody
    • CENF antibody
    • CENP F antibody
    • CENP-F antibody
    • CENPF antibody
    • CENPF kinetochore protein antibody
    • CENPF_HUMAN antibody
    • Centromere protein F 350/400ka antibody
    • Centromere protein F antibody
    • Hcp 1 antibody
    • Hcp1 antibody
    • Kinetochore protein CENP F antibody
    • Kinetochore protein CENPF antibody
    • Mitosin antibody
    • PRO1779 antibody
    see all

Anti-CENPF antibody images

  • ab5 staining CENPF in Mouse embryonic fibroblast cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.3% Triton X-100 and blocked with 3% BSA for 30 minutes at 23°C. Samples were incubated with primary antibody (1/300 in PBS + 0.1% Triton-X 100 + 1% BSA) for 12 hours at 4°C. An Alexa Fluor® 488-conjugated Goat anti-rabbit IgG polyclonal (1/1000) was used as the secondary antibody.

    See Abreview

  • HeLa cells were labelled with anti-ACA and anti-CENPF (ab5). ab5 was used at a working dilution of 1/400. This image demonstrates the dramatic increase in fluorescence that occurs late in G2 cells (indicated by arrows). In the final panel DAPI is pseudo-coloured blue, while ACA and CENPF are coloured green and red respectively. 40x magnification.

  • Interpahse and Prophase HeLa cells were labelled with anti-ACA and anti-CENPF (ab5). ab5 was used at a working dilution of 1/400. This image emphasizes the redistribution of CENPF from the nuclear matrix during late G2 following entry into the initial stages of mitosis (see the accompanying image). Distinct punctate CENPF patterns proximally located in relation to the centromeres can be seen. In the final panel DAPI is pseudo-coloured blue, while ACA and CENPF are green and red respectively. 100x magnification. 

  • ICC/IF image of ab5 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab5, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).

  • All lanes : Anti-CENPF antibody (ab5) at 1/1500 dilution

    Lane 1 : Mitotic
    Lane 2 : Asyn.
    Lane 3 : 50gamma Mitotic HeLa
    Lane 4 : 50gamma Asyn. HeLa lys


    Predicted band size : 330 kDa
    Observed band size : 310 kDa (why is the actual band size different from the predicted?)
  • ab5 staining (human) T98G glioma cells by flow cytometry.  Cells were trypsinized, spun down & resuspended in PBS before fixing and permeabilised in ethanol.  The primary antibody was diluted 1/400 and incubated with the cells for 2 hours at 20°C.  An Alexa Fluor® 488 conjugaetd goat anti-rabit IgG was used as the secondary antibody. 

    Gating strategy: intact cells with >80% and <250% of G1 DNA content

    See Abreview

  • Ab5 staining Human normal colon tissue. Staining is localised to nuclear compartment.
    Left panel: with primary antibody at 4 ug/ml. Right panel: isotype control.
    Sections were stained using an automated system DAKO Autostainer Plus , at room temperature: sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.

References for Anti-CENPF antibody (ab5)

This product has been referenced in:
  • Kruiswijk F  et al. Targeted inhibition of metastatic melanoma through interference with Pin1-FOXM1 signaling. Oncogene N/A:N/A (2015). Read more (PubMed: 26279295) »
  • Carruthers R  et al. Abrogation of radioresistance in glioblastoma stem-like cells by inhibition of ATM kinase. Mol Oncol 9:192-203 (2015). Read more (PubMed: 25205037) »

See all 29 Publications for this product

Product Wall

Application Immunocytochemistry/ Immunofluorescence
Blocking step BSA as blocking agent for 30 minute(s) · Concentration: 3% · Temperature: 23°C
Sample Mouse Cell (Mouse embryonic fibroblast cells)
Specification Mouse embryonic fibroblast cells
Permeabilization Yes - 0.3% Triton X-100
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted Oct 15 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (Human lung adenocarcinoma A549)
Specification Human lung adenocarcinoma A549
Fixative Paraformaldehyde
Permeabilization Yes - 0.2% Triton in PBS
Blocking step BSA as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 20°C
Username

Dr. Stuart Rulten

Verified customer

Submitted Jan 02 2013

Thank you for contacting us.

While I was unable to locate any information regarding the specific protocol used to synchronize the mitotic cells in the image on our website (which was provided by a collaborator), the following protocol may be ...

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (HeLa cells)
Specification HeLa cells
Fixative Formaldehyde
Permeabilization Yes - 0.1% Triton X 100
Blocking step BSA as blocking agent for 30 minute(s) · Concentration: 3% · Temperature: 20°C
Username

Abcam user community

Verified customer

Submitted Jan 29 2008

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Flow Cytometry
Sample Human Cell (T98G glioma cells)
Specification T98G glioma cells
Fixation Ethanol
Permeabilization Yes - Ethanol
Gating Strategy intact cells with >80% and <250% of G1 DNA content
Username

Abcam user community

Verified customer

Submitted Jan 29 2008

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (HeLa)
Specification HeLa
Fixative Paraformaldehyde
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2
Username

Abcam user community

Verified customer

Submitted Aug 15 2006

The exact immunogen is proprietary information which was not provided by the supplier unfortunately. I BLASTed the last 200+ residues (our datasheet says that it was made from a bacterial fusion protein from the C-terminus of CENP-F). It shows ~75% h...

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry
Sample Human Cell (HeLa, T98G)
Specification HeLa, T98G
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1%
Username

Dr. Lesley McPhail

Verified customer

Submitted Nov 09 2005

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (HeLa, SK-N-SH and HCT116)
Specification HeLa, SK-N-SH and HCT116
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted Sep 13 2005

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"