Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-IDH2 antibody [EPR7576] (ab129180)

Overview

  • Product name

    Anti-IDH2 antibody [EPR7576]
    See all IDH2 primary antibodies
  • Description

    Rabbit monoclonal [EPR7576] to IDH2
  • Host species

    Rabbit
  • Tested applications

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

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human IDH2 aa 50-150. The exact sequence is proprietary.

  • Positive control

    • Human thyroid gland carcinoma tissue; Molt-4, K562, 293T and HepG2 whole cell lysate (ab7900).
  • General notes

     

     

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

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab129180 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 - 1/10000. Detects a band of approximately 45 kDa (predicted molecular weight: 51 kDa).
IHC-P 1/250 - 1/500. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Flow Cyt 1/1000 - 1/10000.

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

 

  • Application notes
    Is unsuitable for ICC/IF or IP.
  • Target

    • Function

      Plays a role in intermediary metabolism and energy production. It may tightly associate or interact with the pyruvate dehydrogenase complex.
    • Involvement in disease

      D-2-hydroxyglutaric aciduria 2
      Glioma
      enetic variations are associated with cartilaginous tumors such as enchondroma or chondrosarcoma.
    • Sequence similarities

      Belongs to the isocitrate and isopropylmalate dehydrogenases family.
    • Post-translational
      modifications

      Acetylation at Lys-413 dramatically reduces catalytic activity. Deacetylated by SIRT3.
    • Cellular localization

      Mitochondrion.
    • Information by UniProt
    • Database links

    • Alternative names

      • D2HGA2 antibody
      • ICD-M antibody
      • IDH antibody
      • IDH2 antibody
      • IDHM antibody
      • IDHP_HUMAN antibody
      • IDP antibody
      • IDPM antibody
      • Isocitrate dehydrogenase [NADP], mitochondrial antibody
      • Isocitrate dehydrogenase 2 (NADP+), mitochondrial antibody
      • mNADP-IDH antibody
      • NADP(+)-specific ICDH antibody
      • Oxalosuccinate decarboxylase antibody
      see all

    Images

    • Lane 1: Wild-type HAP1 cell lysate (20 µg)
      Lane 2: IDH2 knockout HAP1 cell lysate (20 µg)
      Lane 3: K562 cell lysate (20 µg)
      Lane 4: HepG2 cell lysate (20 µg)
      Lanes 1 - 4: Merged signal (red and green). Green - ab129180 observed at 47 kDa. Red - loading control, ab8245, observed at 37 kDa.
      ab129180 was shown to recognize IDH2 when IDH2 knockout samples were used, along with additional cross-reactive bands. Wild-type and IDH2 knockout samples were subjected to SDS-PAGE. ab129180 and ab8245 (loading control to GAPDH) were diluted 1/1000 and 1/2000 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.

    • ab129180, at 1/250 dilution, staining IDH2 in Formalin-fixed, Paraffin-embedded Human thyroid gland carcinoma by Immunohistochemistry.

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    • Overlay histogram showing MCF7 cells stained with ab129180 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab129180, 1/10000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in MCF7 cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
    • All lanes : Anti-IDH2 antibody [EPR7576] (ab129180) at 1/1000 dilution

      Lane 1 : Molt-4 cell lysate
      Lane 2 : K562 cell lysate
      Lane 3 : 293T cell lysate
      Lane 4 : HepG2 cell lysate

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Goat Anti-rabbit HRP

      Predicted band size: 51 kDa
      Observed band size: 45 kDa
      why is the actual band size different from the predicted?

    References

    This product has been referenced in:

    • Benlamara S  et al. Distinctive Krebs cycle remodeling in iPSC-derived neural and mesenchymal stem cells. Biochem Biophys Res Commun 511:658-664 (2019). Read more (PubMed: 30826061) »
    • Pichitpunpong C  et al. Phenotypic subgrouping and multi-omics analyses reveal reduced diazepam-binding inhibitor (DBI) protein levels in autism spectrum disorder with severe language impairment. PLoS One 14:e0214198 (2019). Read more (PubMed: 30921354) »
    See all 5 Publications for this product

    Customer reviews and Q&As

    1-4 of 4 Abreviews or Q&A

    Application
    Western blot
    Sample
    Rat Tissue lysate - whole (Retina, RPE)
    Gel Running Conditions
    Reduced Denaturing (12% gel)
    Loading amount
    20 µg
    Specification
    Retina, RPE
    Blocking step
    Licor Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C

    Abcam user community

    Verified customer

    Submitted Jul 25 2017

    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (Human ARPE-19 cells)
    Gel Running Conditions
    Reduced Denaturing (12%)
    Loading amount
    20 µg
    Specification
    Human ARPE-19 cells
    Blocking step
    Licor Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C

    Abcam user community

    Verified customer

    Submitted Aug 23 2016

    Application
    Western blot
    Sample
    Mouse Tissue lysate - whole (Brain, Heart, Liver, Muscle)
    Gel Running Conditions
    Reduced Denaturing (10%, hand-cast)
    Loading amount
    20 µg
    Specification
    Brain, Heart, Liver, Muscle
    Blocking step
    Licor Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C

    Abcam user community

    Verified customer

    Submitted Jul 20 2016

    Answer

    Thank you and your customer for your inquiry.

    Please find the recommended antigen retrieval procedure for both antibodies below:

    Antigen Retrieval


    This is recommended Heat Induced Epitope Retrieval (HIER) using Decloaking Chamber/Pressure Cooker. Hot water bath or Microwave with temperature sensor can be also used (protocol would vary depending on the method used).

    1. Add 500 ml of dH 2O to Decloaker/Pressure Cooker.

    2. Immerse slides into staining dish containing Antigen Retrieval Solution: 0.01M Sodium Citrate Buffer, pH 6.0. Place staining dish into decloaking chamber.

    3. Program to run for 30 seconds at 125° C, followed by 10 seconds at 90° C.

    4. Let it cool down to room temperature (10 - 20 minutes).

    5. Removes slides and rinse in TBST.

    6. Proceed to Staining step.

    The dilution might have to be optimised individually, but I can confirm that we had good results with 1/250 for ab129180 with Human thyroid gland carcinoma and 1/50 for ab117976 with Human breast tissue, colon tissue and endometrium tissue.

    I hope this information is helpful and wish you customer good luck with his research.

    Read More

    Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
    For licensing inquiries, please contact partnerships@abcam.com

    Sign up