Validated using a knockout cell line
Recombinant
RabMAb

Anti-Glutaminase antibody [EP7212] (ab156876)

Overview

  • Product name
    Anti-Glutaminase antibody [EP7212]
    See all Glutaminase primary antibodies
  • Description
    Rabbit monoclonal [EP7212] to Glutaminase
  • Host species
    Rabbit
  • Tested applications
    Suitable for: Flow Cyt, WB, IHC-P, ICC/IFmore details
  • Species reactivity
    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human Glutaminase aa 50-150. The exact sequence is proprietary.
    (Peptide available as ab206976)

  • Positive control
    • WB: Human fetal brain and fetal kidney brain tissue lysates, HeLa and 293T cell lysates. IHC-P: Human tonsil, kidney, brain, glioma and pancreas tissues. ICC/IF: HeLa cells. Flow Cyt: HeLa cells.
  • 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

    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.

Properties

Applications

Our Abpromise guarantee covers the use of ab156876 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
Flow Cyt 1/400.
WB 1/1000 - 1/10000. Predicted molecular weight: 73 kDa.Can be blocked with Human Glutaminase peptide (ab206976).
IHC-P 1/100 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

ICC/IF 1/250 - 1/500.

Target

  • Function
    Catalyzes the first reaction in the primary pathway for the renal catabolism of glutamine.
  • Tissue specificity
    KGA is expressed predominantly in brain and kidney but not in liver, isoform 3 is expressed principally in cardiac muscle and pancreas but not in liver or brain, and isoform 2 is expressed solely in cardiac and skeletal muscle.
  • Sequence similarities
    Belongs to the glutaminase family.
    Contains 1 ANK repeat.
  • Cellular localization
    Mitochondrion.
  • Information by UniProt
  • Database links
  • Alternative names
    • AAD20 antibody
    • DKFZp686O15119 antibody
    • FLJ10358 antibody
    • GAC antibody
    • GAM antibody
    • GLS antibody
    • GLS1 antibody
    • GLSK_HUMAN antibody
    • Glutaminase C antibody
    • Glutaminase kidney isoform antibody
    • Glutaminase phosphate activated antibody
    • K-glutaminase antibody
    • KGA antibody
    • KIAA0838 antibody
    • L-glutamine amidohydrolase antibody
    • mitochondrial antibody
    see all

Images

  • Lane 1: Wild type HAP1 whole cell lysate (20 µg)
    Lane 2: GLS knockout HAP1 whole cell lysate (20 µg)
    Lane 3: HeLa whole cell lysate (20 µg)
    Lane 4: HEK293 whole cell lysate (20 µg)
    Lanes 1 - 4: Merged signal (red and green). Green - ab156876 observed at 62 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab156876 was shown to specifically recognize GLS in wild-type HAP1 cells along with additional cross reactive bands. No bands were observed when GLS knockout samples were used. Wild-type and GLS knockout samples were subjected to SDS-PAGE. Ab156876 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/10,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1hr at room temperature before imaging.

  • Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling Glutaminase with purified ab156876 at 1/250. Cells were fixed with 100% methanol and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) were also used.

    Control 1: primary antibody (1/250) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000).

    Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000).

  • Flow cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling Glutaminase (red) with ab156876 at a 1/400 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (ab172730). Blue (unlabeled control) - Cells without incubation with the primary and secondary antibodies.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cerebral cortex tissue labelling Glutaminase with purified ab156876 at a dilution of 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • All lanes : Anti-Glutaminase antibody [EP7212] (ab156876) at 10 µg (purified)

    Lane 1 : Human fetal kidney tissue lysate
    Lane 2 : HeLa whole cell lysate
    Lane 3 : HEK293 whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 73 kDa
    Observed band size: 65,73 kDa (why is the actual band size different from the predicted?)



    Blocking and dilution buffer: 5% NFDM /TBST.

    Recognizes endogenous levels of total Glutaminase protein (both the 73 kDa KGA and 65 kDa GAC isoforms).

  • Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling Glutaminase with unpurified ab156876 antibody at a dilution of 1/250.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human glioma tissue labelling Glutaminase with purified ab156876 at a dilution of 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • All lanes : Anti-Glutaminase antibody [EP7212] (ab156876) at 1/1000 dilution (unpurified)

    Lane 1 : Human fetal brain extract
    Lane 2 : Human fetal kidney extract
    Lane 3 : HeLa cell lysate
    Lane 4 : 293T cell lysate

    Lysates/proteins at 10 µg per lane.

    Developed using the ECL technique.

    Predicted band size: 73 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of normal human tonsil tissue labelling Glutaminase with unpurified ab156876.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of normal human brain tissue labelling Glutaminase with unpurified ab156876.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human kidney tissue labelling Glutaminase with unpurified ab156876 at a dilution of 1/100.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human pancreas tissue labelling Glutaminase with unpurified ab156876 at a dilution of 1/100.

  • Unpurified ab156876 showing negative staining in Human skeletal muscle.

  • Unpurified ab156876 showing negative staining in Human heart.

References

This product has been referenced in:
  • Lynch DS  et al. GLS loss of function causes autosomal recessive spastic ataxia and optic atrophy. Ann Clin Transl Neurol 5:216-221 (2018). Read more (PubMed: 29468182) »
  • Duan G  et al. Increased Glutamine Consumption in Cisplatin-Resistant Cells Has a Negative Impact on Cell Growth. Sci Rep 8:4067 (2018). Read more (PubMed: 29511244) »

See all 9 Publications for this product

Customer reviews and Q&As

The actual band size detected by ab156876 (65 kDa) is lower than the expected size (73 kDa) which could be due to the fact that the antibody can detect all three isoforms. You may wish to look at this site for further information about the different is...

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Application
Western blot
Loading amount
10 µg
Gel Running Conditions
Reduced Denaturing (7.5% SDS-PAGE)
Sample
Human Cell lysate - whole cell (Breast Cancer Cells)
Specification
Breast Cancer Cells
Treatment
10uM Compounds A and B for 24hrs
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Username

Karina Ricart

Verified customer

Submitted Aug 08 2014

ab156876: Based on the immunogen sequence, this antibody is capable of detecting mouse kidney GLS1 isoforms 1 and 2 according to swissprot:

mouse GLS1: http://www.uniprot.org/uniprot/D3Z7P3

ab113509: I blasted human GLS2 against mouse...

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