Overview

  • Product name
    Anti-GAPDH antibody [EPR6256] - BSA and Azide free
    See all GAPDH primary antibodies
  • Description
    Rabbit monoclonal [EPR6256] to GAPDH - BSA and Azide free
  • Host species
    Rabbit
  • Specificity

    The African green monkey recommendation is based on the WB results. We do not guarantee IHC-P for African green monkey.

  • Tested applications
    Suitable for: WB, ICC/IF, IP, IHC-P, Flow Cytmore details
  • Species reactivity
    Reacts with: Human, African green monkey
  • Immunogen

    Synthetic peptide corresponding to a region within Human GAPDH (Uniprot: P04406)

  • Positive control
    • WB: 293T, HeLa, HepG2, HUVEC, MCF7 or SH SY5Y cell lysates. ICC/IF: MCF7 and HeLa cells IHC-P: Human bladder carcinoma FC: HeLa cells IP: HeLa cell lysate
  • General notes

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

    The formulation and the concentration of this product is compatible for metal-conjugation for mass cytometry (CyTOF®).

    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.

    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 ab186930 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: 36 kDa.

Please check the parent abID, ab128915, for a recommended dilution.

ICC/IF Use at an assay dependent concentration.
IP Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration.

See IHC antigen retrieval protocols.

The African green monkey recommendation is based on the WB results. We do not guarantee IHC-P for African green monkey.

Flow Cyt Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

Target

  • Function
    Has both glyceraldehyde-3-phosphate dehydrogenase and nitrosylase activities, thereby playing a role in glycolysis and nuclear functions, respectively. Participates in nuclear events including transcription, RNA transport, DNA replication and apoptosis. Nuclear functions are probably due to the nitrosylase activity that mediates cysteine S-nitrosylation of nuclear target proteins such as SIRT1, HDAC2 and PRKDC (By similarity). Glyceraldehyde-3-phosphate dehydrogenase is a key enzyme in glycolysis that catalyzes the first step of the pathway by converting D-glyceraldehyde 3-phosphate (G3P) into 3-phospho-D-glyceroyl phosphate.
  • Pathway
    Carbohydrate degradation; glycolysis; pyruvate from D-glyceraldehyde 3-phosphate: step 1/5.
  • Sequence similarities
    Belongs to the glyceraldehyde-3-phosphate dehydrogenase family.
  • Post-translational
    modifications
    S-nitrosylation of Cys-152 leads to interaction with SIAH1, followed by translocation to the nucleus.
    ISGylated.
  • Cellular localization
    Cytoplasm > cytosol. Nucleus. Cytoplasm > perinuclear region. Membrane. Translocates to the nucleus following S-nitrosylation and interaction with SIAH1, which contains a nuclear localization signal (By similarity). Postnuclear and Perinuclear regions.
  • Information by UniProt
  • Database links
  • Alternative names
    • 38 kDa BFA-dependent ADP-ribosylation substrate antibody
    • aging associated gene 9 protein antibody
    • Aging-associated gene 9 protein antibody
    • BARS-38 antibody
    • cb609 antibody
    • EC 1.2.1.12 antibody
    • Epididymis secretory sperm binding protein Li 162eP antibody
    • G3P_HUMAN antibody
    • G3PD antibody
    • G3PDH antibody
    • GAPD antibody
    • GAPDH antibody
    • Glyceraldehyde 3 phosphate dehydrogenase antibody
    • Glyceraldehyde-3-phosphate dehydrogenase antibody
    • HEL-S-162eP antibody
    • KNC-NDS6 antibody
    • MGC102544 antibody
    • MGC102546 antibody
    • MGC103190 antibody
    • MGC103191 antibody
    • MGC105239 antibody
    • MGC127711 antibody
    • MGC88685 antibody
    • OCAS, p38 component antibody
    • OCT1 coactivator in S phase, 38-KD component antibody
    • peptidyl cysteine S nitrosylase GAPDH antibody
    • Peptidyl-cysteine S-nitrosylase GAPDH antibody
    • wu:fb33a10 antibody
    see all

Images

  • Anti-GAPDH antibody [EPR6256] - BSA and Azide free (ab186930) (unpurified) + HEK293 (human embryonic kidney) whole cell lysate at 10 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051)

    Predicted band size: 36 kDa


    Exposure time: 30 seconds


    Blocking buffer and concentration: 5% NFDM/TBST

    Diluting buffer and concentration: 5% NFDM/TBST

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human bladder carcinoma tissue sections labeling GAPDH with purified ab128915 at 1/2000 dilution (0.06 µg/ml). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab128915).

  • Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling GAPDH with purified ab128915 at 1/250 dilution (0.4 µg/ml). Cells were fixed in 100% Methanol. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200 (2.5 µg/ml) dilution. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab128915).

  • Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling GAPDH with purified ab128915 at 1/20 dilution (10 µg/ml) (red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).
    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab128915).

  • ab128915 (purified) at 1/20 dilution (0.5ug) immunoprecipitating GAPDH in HeLa whole cell lysates.
    Lane 1: HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates 10ug
    Lane 2 (+): ab128915 & HeLa whole cell lysates
    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab128915 in HeLa whole cell lysates
    For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.
    Blocking and diluting buffer: 5% NFDM/TBST.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab128915).

  • IHC image of ab128915 (unpurified) staining GAPDH in human pancreas* formalin fixed paraffin embedded tissue sections, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab128915, 1:250 dilution, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. No primary antibody was used in the secondary only control (shown on the inset).
    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab128915).

  • ab128915 (unpurified) staining GAPDH in HeLa (human epithelial cell line from cervix adenocarcinoma) cells. The cells were fixed with 100% methanol (5min) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated with ab128915 at 2μg/ml and ab7291 at 1µg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with an goat anti-rabbit AlexaFluor®488 (ab150081) at 2 μg/ml (shown in green) and goat anti-mouse AlexaFluor®594 (ab150120) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labelled in blue with DAPI.

    Negative controls: 1– Rabbit primary antibody and anti-mouse secondary antibody; 2 – Mouse primary antibody and anti-rabbit secondary antibody. Controls 1 and 2 indicate that there is no unspecific reaction between primary and secondary antibodies used.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab128915).

  • ab128915 (unpurified) staining GAPDH in human HeLa (human epithelial cell line from cervix adenocarcinoma) cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde and permeabilized with 0.5% Triton X-100 in PBS. Samples were incubated with primary antibody (1/500 in PBS) for 1 hour at 22°C. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG polyclonal (1/200) was used as the secondary antibody. Counter stained with DAPI.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab128915).

  • Overlay histogram showing HeLa (human epithelial cell line from cervix adenocarcinoma) cells stained with ab128915 (unpurified) (red line). The cells were fixed with 80% methanol (5 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 (ab128915, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was goat anti-rabbit Alexa Fluor® 488 (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 data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab128915).

  • ab128915 (unpurified), at 1/250, staining GAPDH in MCF7 (human breast adenocarcinoma cell line) cells by immunofluorescence.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab128915).

  • Anti-GAPDH antibody [EPR6256] - BSA and Azide free (ab186930) (unpurified) + HeLa (human cervix adenocarcinoma) whole cell lysate at 10 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051)

    Predicted band size: 36 kDa


    Exposure time: 1 minute


    Blocking buffer and concentration: 5% NFDM/TBST

    Diluting buffer and concentration: 5% NFDM/TBST

References

This product has been referenced in:
  • Rossetto CC  et al. Cis and trans acting factors involved in human cytomegalovirus experimental and natural latent infection of CD14 (+) monocytes and CD34 (+) cells. PLoS Pathog 9:e1003366 (2013). CLIP ; Human . Read more (PubMed: 23717203) »
  • Zhang J  et al. Gankyrin plays an essential role in estrogen-driven and GPR30-mediated endometrial carcinoma cell proliferation via the PTEN/PI3K/AKT signaling pathway. Cancer Lett 339:279-87 (2013). Read more (PubMed: 23142288) »
See all 3 Publications for this product

Customer reviews and Q&As

There are currently no Customer reviews or Questions for ab186930.
Please use the links above to contact us or submit feedback about this product.

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