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Cell Biology Cell Cycle Kinases/Phosphatases Phosphatases
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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)

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  • Certificate of Compliance
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Western blot - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
  • Immunocytochemistry/ Immunofluorescence - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
  • Immunoprecipitation - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
  • Flow Cytometry - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
  • Western blot - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
  • Flow Cytometry - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
  • Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [E302] to Cdc25C - BSA and Azide free
  • Suitable for: WB, IP, IHC-P, ICC/IF, Flow Cyt
  • Knockout validated
  • Reacts with: Human

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Overview

  • Product name

    Anti-Cdc25C antibody [E302] - BSA and Azide free
    See all Cdc25C primary antibodies
  • Description

    Rabbit monoclonal [E302] to Cdc25C - BSA and Azide free
  • Host species

    Rabbit
  • Specificity

    The antibody can also detect splice isoform 2, 4 and 5 of human Cdc25C, based on sequence homology.
  • Tested applications

    Suitable for: WB, IP, IHC-P, ICC/IF, Flow Cytmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human Cdc25C aa 1-100 (N terminal). The exact sequence is proprietary.

  • Positive control

    • WB: HAP1 and HeLa cell lysate. IHC-P: Human pancreas and urinary bladder tissues. IP: HeLa cell lysate. Flow Cyt: K562 and HeLa cells. IHC-P: HeLa cells.
  • General notes

    ab232553 is the carrier-free version of ab32444 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.

    Ab232553 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

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

    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
    For more information see here.

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

    Reproducibility is key to advancing scientific discovery and accelerating scientists’ next breakthrough.

    Abcam is leading the way with our range of recombinant antibodies, knockout-validated antibodies and knockout cell lines, all of which support improved reproducibility.

    We are also planning to innovate the way in which we present recommended applications and species on our product datasheets, so that only applications & species that have been tested in our own labs, our suppliers or by selected trusted collaborators are covered by our Abpromise™ guarantee.

    In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.

    We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.

    Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.

    Please check that this product meets your needs before purchasing. If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, as well as customer reviews and Q&As.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C. Do Not Freeze.
  • Storage buffer

    pH: 7.2
    Constituent: PBS
  • Carrier free

    Yes
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    E302
  • Isotype

    IgG
  • Research areas

    • Cell Biology
    • Cell Cycle
    • Kinases/Phosphatases
    • Phosphatases
    • Epigenetics and Nuclear Signaling
    • Cell cycle
    • Kinases/Phosphatases
    • Phospatases

Associated products

  • Alternative Versions

    • Alexa Fluor® 488 Anti-Cdc25C antibody [E302] (ab205425)
    • Alexa Fluor® 647 Anti-Cdc25C antibody [E302] (ab205426)
    • HRP Anti-Cdc25C antibody [E302] (ab205734)
    • Anti-Cdc25C antibody [E302] (ab32444)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
    • Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773)
  • Conjugation kits

    • APC Conjugation Kit - Lightning-Link® (ab201807)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • KO cell lines

    • Human CDC25C knockout HeLa cell line (ab265189)
  • KO cell lysates

    • Human CDC25C knockout HeLa cell lysate (ab257387)
  • Recombinant Protein

    • Recombinant Human Cdc25C protein (ab158076)

Applications

Our Abpromise guarantee covers the use of ab232553 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: 53 kDa.
IP Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.

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.
  • Target information above from: UniProt accession P30307 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 995 Human
    • Omim: 157680 Human
    • SwissProt: P30307 Human
    • Unigene: 656 Human
    • 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

    • Western blot - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      Western blot - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      All lanes : Anti-Cdc25C antibody [E302] (ab32444) at 1/1000 dilution

      Lane 1 : Wild-type HeLa cell lysate
      Lane 2 : CDC25C knockout HeLa cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 53 kDa
      Observed band size: 58 kDa
      why is the actual band size different from the predicted?



      This data was developed using the same antibody clone in a different buffer formulation (ab32444).

        Lanes 1- 2: Merged signal (red and green). Green - ab32444 observed at 58 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

       ab32444 was shown to react with Cdc25C in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab265189 (knockout cell lysate ab257387) was used. Wild-type HeLa and CDC25C knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab32444 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)

      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.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32444).

    • Immunocytochemistry/ Immunofluorescence - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      Immunocytochemistry/ Immunofluorescence - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)

      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). 

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32444).

    • Immunoprecipitation - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      Immunoprecipitation - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)

      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, VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

      Blocking buffer and concentration: 5% NFDM/TBST.

      Diluting buffer and concentration: 5% NFDM /TBST.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32444).

    • Flow Cytometry - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      Flow Cytometry - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)

      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.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32444).

    • Western blot - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      Western blot - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)

      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: Human 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 1/2,500 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 1/10,000 dilution for 1 hour at room temperature before imaging.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32444).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)

      Immunohistochemical analysis of paraffin-embedded human urinary bladder carcinoma unpurified ab32444 at 1/250 dilution.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32444).

    • Flow Cytometry - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      Flow Cytometry - Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)This image is courtesy of an Abreview submitted by Brandon White.

      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.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32444).

    • Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)
      Anti-Cdc25C antibody [E302] - BSA and Azide free (ab232553)

    Protocols

    To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

    Click here to view the general protocols

    Datasheets and documents

    • Datasheet

    Certificate of Compliance

    To download a Certificate of Compliance, please enter your Lot number below:

  • References (0)

    Publishing research using ab232553? Please let us know so that we can cite the reference in this datasheet.

    ab232553 has not yet been referenced specifically in any publications.

    Customer reviews and Q&As

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