Astana Biomed Group, an authorized Abcam distributor in Central Asia

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

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 & Species

    Application Species
    Flow Cyt
    Human
    ICC/IF
    Human
    IHC-P
    Human
    IP
    Human
    WB
    Human
    See all applications and species data

  • Immunogen

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • 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

    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.

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing the problem with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation.

    One factor contributing to the crisis is the use of antibodies that are not suitable. This can lead to misleading results and the use of incorrect data informing project assumptions and direction. To help address this challenge, we have introduced an application and species grid on our primary antibody datasheets to make it easy to simplify identification of the right antibody for your needs.

    Learn more here.

Properties

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)

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