Overview

  • Product name
    Anti-Cdc25C antibody [E302]
    See all Cdc25C primary antibodies
  • Description
    Rabbit monoclonal [E302] to Cdc25C
  • Specificity
    The antibody can also detect splice isoform 2, 4 and 5 of human Cdc25C, based on sequence homology.
  • Tested applications
    Suitable for: WB, IHC-P, ICC/IF, Flow Cyt, IPmore details
  • Species reactivity
    Reacts with: Human
  • Immunogen

    A synthetic peptide corresponding to residues near N-terminus of human Cdc25C.

  • Positive control
    • HeLa cell lysate and human urinary bladder carcinoma tissue.
  • General notes

    Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.

     

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

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab32444 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 1/1000 - 1/5000. Detects a band of approximately 60 kDa (predicted molecular weight: 53 kDa).
IHC-P 1/2500.

For unpurified, use 1/250 - 1/500.

ICC/IF 1/250 - 1/500.
Flow Cyt 1/180.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

IP 1/30 - 1/80.

Target

  • Function
    Functions as a dosage-dependent inducer in mitotic control. It is a tyrosine protein phosphatase required for progression of the cell cycle. It directly dephosphorylates CDK1 and activate its kinase activity.
  • Sequence similarities
    Belongs to the MPI phosphatase family.
    Contains 1 rhodanese domain.
  • Developmental stage
    Expressed predominantly in G2 phase.
  • Post-translational
    modifications
    Phosphorylated by CHK1 on Ser-216. This phosphorylation creates a binding site for 14-3-3 protein and inhibits the phosphatase. Phosphorylated by PLK4.
  • Cellular localization
    Nucleus.
  • Information by UniProt
  • Database links
  • Alternative names
    • CDC 25 antibody
    • Cdc 25C antibody
    • CDC25 antibody
    • CDC25C antibody
    • Cell division cycle 25 homolog C antibody
    • Cell division cycle 25C antibody
    • Cell division cycle 25C protein antibody
    • Dual specificity phosphatase Cdc25C antibody
    • M phase inducer phosphatase 3 antibody
    • M-phase inducer phosphatase 3 antibody
    • Mitosis inducer CDC25 antibody
    • MPIP3 antibody
    • MPIP3_HUMAN antibody
    • Phosphotyrosine phosphatase antibody
    • PPP1R60 antibody
    • protein phosphatase 1, regulatory subunit 60 antibody
    see all

Images

  • Anti-Cdc25C antibody [E302] (ab32444) at 1/1000 dilution (purified) + HeLa (human cervix adenocarcinoma) whole cell lysate at 20 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size : 53 kDa
    Observed band size : 60 kDa (why is the actual band size different from the predicted?)

    Blocking and diluting buffer 5% NFDM/TBST

  • Immunohistochemical analysis of paraffin embedded human pancreas tissue section labelling Cdc25C with purified ab32444 at dilution of 1/2500. The secondary antibody used was Goat Anti-Rabbit IgG H&L (HRP) (ab97051), at dilution of 1/500. The sample was counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

  • Immunocytochemistry/Immunofluorescence analysis of HeLa (human cervix adenocarcinoma) cells labelling Cdc25C with purified ab32444 at 1/400. Cells were fixed with 4% Paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. The cells were co-stained with ab7291, a mouse anti-tubulin antibody (1/1000) using ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) as the secondary. Nuclei couterstained with DAPI (blue).

    For negative control 1, rabbit primary antibody was used, followed by anti-mouse secondary antibody (ab150120). For negative control 2, mouse primary antibody (ab7291) was used followed by anti-rabbit secondary antibody (ab150077). 

  • Flow Cytometry analysis of K562 (human chronic myelogenous leukemia) cells labelling Cdc25C with purified ab32444 at 1/180 (red). Cells were fixed with 4% paraformaldehyde. Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

  • Ab32444 (purified) at 1/30 immunoprecipitating Cdc25C in HeLa (human cervix adenocarcinoma) whole cell lysate.

    Lane 1 (input): HeLa (human cervix adenocarcinoma) whole cell lysate

    Lane 2 (+): ab32444 + HeLa (human cervix adenocarcinoma) whole cell lysate

    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32444 in HeLa (human cervix adenocarcinoma) whole cell lysate

    For western blotting, ab131366 VeriBlot for IP (HRP) was used as the secondary antibody (1/10000).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.



  • Predicted band size : 53 kDa

    Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: Cdc25C knockout HAP1 cell lysate (20 µg)
    Lane 3: HeLa cell lysate (20 µg)
    Lane 4: Hu bladder cell lysate (20 µg)
    Lanes 1 - 4: Merged signal (red and green). Green - ab32444 observed at 55 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab32444 was shown to recognize Cdc25C when Cdc25C knockout samples were used, along with additional cross-reactive bands. Wild-type and Cdc25C knockout samples were subjected to SDS-PAGE. ab32444 and ab8245 (loading control to GAPDH) were diluted at 1/2500 and 1/10 000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.

  • All lanes : Anti-Cdc25C antibody [E302] (ab32444) at 1/5000 dilution (purified)

    Lane 1 : K562 (human chronic myelogenous leukemia) whole cell lysate
    Lane 2 : HEK293 (human embryonic kidney) whole cell lysates

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size : 53 kDa
    Observed band size : 60 kDa (why is the actual band size different from the predicted?)

    Blocking and diluting buffer 5% NFDM/TBST

  • Anti-Cdc25C antibody [E302] (ab32444) at 1/5000 dilution (unpurified) + HeLa cell lysate

    Predicted band size : 53 kDa
    Observed band size : 60 kDa (why is the actual band size different from the predicted?)
  • Immunohistochemical analysis of paraffin-embedded human urinary bladder carcinoma unpurified ab32444 at 1/250 dilution.

    Immunohistochemical analysis of paraffin-embedded human urinary bladder carcinoma ab32444 at 1/250 dilution.
  • Flow cytometry analysis of HeLa cells, staining Cdc25C with unpurified ab32444.

    Cells were fixed with formaldehyde and permeabilized with 90% methanol. Samples were incubated with primary antibody (1/20 in PBS + 10% goat serum) for 1 hour at 23°C. A FITC-conjugated goat anti-rabbit polyclonal IgG (1/1000) was used as the secondary antibody.

    See Abreview

References

This product has been referenced in:
  • Skowron KB  et al. Basal Tumor Cell Isolation and Patient-Derived Xenograft Engraftment Identify High-Risk Clinical Bladder Cancers. Sci Rep 6:35854 (2016). WB, IHC ; Human . Read more (PubMed: 27775025) »
  • Zhu X  et al. Ziyuglycoside I Inhibits the Proliferation of MDA-MB-231 Breast Carcinoma Cells through Inducing p53-Mediated G2/M Cell Cycle Arrest and Intrinsic/Extrinsic Apoptosis. Int J Mol Sci 17:N/A (2016). Human . Read more (PubMed: 27879682) »

See all 6 Publications for this product

Customer reviews and Q&As

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (HeLa cell lysate)
Loading amount
20 µg
Specification
HeLa cell lysate
Gel Running Conditions
Reduced Denaturing (4-12% Bis-Tris)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted May 15 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (Human Fibroblast)
Loading amount
15 µg
Specification
Human Fibroblast
Gel Running Conditions
Reduced Denaturing (4-12% Bis-Tris Nupage gel)
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Feb 25 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Flow Cytometry
Sample
Human Cell (HeLa Cells)
Specification
HeLa Cells
Preparation
Cell harvesting/tissue preparation method: Trypsin/EDTA
Sample buffer: Complete Media then PBS
Fixation
Formaldehyde
Permeabilization
Yes - 90% Methanol
Username

Dr. Brandon White

Verified customer

Submitted Aug 30 2012

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

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