Overview

  • Product name
    Anti-MGMT antibody [MT3.1]
    See all MGMT primary antibodies
  • Description
    Mouse monoclonal [MT3.1] to MGMT
  • Host species
    Mouse
  • Tested applications
    Suitable for: IHC-Fr, WB, IHC-FoFr, ICC, Flow Cyt, ICC/IF, IHC-Pmore details
  • Species reactivity
    Reacts with: Human
  • Immunogen

    Recombinant full length protein corresponding to Human MGMT.

  • Positive control
    • Tonsil

Properties

Applications

Our Abpromise guarantee covers the use of ab39253 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
IHC-Fr 1/50.
WB 1/500. Detects a band of approximately 21 kDa.
IHC-FoFr 1/25 - 1/50.
ICC Use at an assay dependent concentration.
Flow Cyt 1/50.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

 

ICC/IF Use at an assay dependent concentration.
IHC-P 1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Target

  • Function
    Involved in the cellular defense against the biological effects of O6-methylguanine (O6-MeG) in DNA. Repairs alkylated guanine in DNA by stoichiometrically transferring the alkyl group at the O-6 position to a cysteine residue in the enzyme. This is a suicide reaction: the enzyme is irreversibly inactivated.
  • Sequence similarities
    Belongs to the MGMT family.
  • Cellular localization
    Nucleus.
  • Information by UniProt
  • Database links
  • Alternative names
    • 6 O methylguanine DNA methyltransferase antibody
    • 6-O-methylguanine-DNA methyltransferase antibody
    • Agat antibody
    • AGT antibody
    • AI267024 antibody
    • EC 2.1.1.63 antibody
    • Methylated DNA protein cysteine methyltransferase antibody
    • Methylated-DNA--protein-cysteine methyltransferase antibody
    • Methylguanine DNA methyltransferase antibody
    • MGC107020 antibody
    • MGMT antibody
    • MGMT_HUMAN antibody
    • O 6 methylguanine DNA alkyltransferase antibody
    • O 6 methylguanine DNA methyltransferase antibody
    • O-6-methylguanine-DNA methyltransferase antibody
    • O-6-methylguanine-DNA-alkyltransferase antibody
    see all

Images

  • Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: MGMT knockout HAP1 cell lysate (20 µg)
    Lane 3: MCF7 cell lysate (20 µg)
    Lane 4: Jurkat cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab39253 observed at 22 kDa. Red - loading control, ab181602, observed at 37 kDa.
    ab39253 was shown to specifically react with MGMT when MGMT knockout samples were used. Wild-type and MGMT knockout samples were subjected to SDS-PAGE. ab39253 and ab181602 (loading control to GAPDH) were diluted 1/500 and 1/10 000 and incubated overnight at 4°C. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1/10 000 dilution for 1 h at room temperature before imaging.

  • ab39253 staining MGMT in Human stomach tissue sections by IHC-Fr (frozen sections). Tissue was fixed with paraformaldehyde and blocked with 5% serum for 1 hour at 23°C. Samples were incubated with primary antibody (1/50 in blocking buffer) for 1 hour at 23°C. An undiluted HRP-conjugated goat anti-mouse IgG polyclonal was used as secondary antibody.

    See Abreview

  • ab39253 staining MGMT in human stomach tissue sections by Immunohistochemistry. Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using 10mM citrate buffer pH6.0. Samples were then blocked with 5% serum for 1 hour at 23°C followed by incubation with the primary antibody, at a 1/100 dilution, for 1 hour at 23°C. An undiluted HRP-conjugated goat anti-mouse IgG polyclonal was used as secondary antibody.

    See Abreview

  • Overlay histogram showing Jurkat cells stained with ab39253 (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 (ab39253, 1/50 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

  • All lanes : Anti-MGMT antibody [MT3.1] (ab39253) at 1/500 dilution (for 16 hours at 4°C)

    All lanes : Human OE33 cells - whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Rabbit Anti-Mouse IgG H&L (HRP) (ab6728) at 1/1000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Observed band size: 21 kDa
    why is the actual band size different from the predicted?


    Exposure time: 30 seconds


    Blocking Step: 5% BSA for 1 hour at 23°C

    See Abreview

  • Immunohistochemical analysis of formalin-fixed, paraffin-embedded Human tonsil tissue, staining MGMT with ab39253.

References

This product has been referenced in:
  • Zink LM  et al. H3.Y discriminates between HIRA and DAXX chaperone complexes and reveals unexpected insights into human DAXX-H3.3-H4 binding and deposition requirements. Nucleic Acids Res 45:5691-5706 (2017). Read more (PubMed: 28334823) »
  • Wang K  et al. Hedgehog/Gli1 signaling pathway regulates MGMT expression and chemoresistance to temozolomide in human glioblastoma. Cancer Cell Int 17:117 (2017). Read more (PubMed: 29225516) »
See all 9 Publications for this product

Customer reviews and Q&As

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1-6 of 6 Abreviews

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HeLa)
Permeabilization
Yes - 0.5% Triton X-100
Specification
HeLa
Fixative
Paraformaldehyde

Dr. Kirk Mcmanus

Verified customer

Submitted Jul 03 2018

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - other (OE33 cells)
Loading amount
20 µg
Specification
OE33 cells
Gel Running Conditions
Reduced Denaturing
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C

Abcam user community

Verified customer

Submitted Aug 27 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (AGS Gastric Carcinoma cells)
Specification
AGS Gastric Carcinoma cells
Fixative
Formaldehyde
Permeabilization
Yes - 0.025% Triton-X in TBS
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C

Abcam user community

Verified customer

Submitted Jun 02 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry
Sample
Human Cultured Cells (AGS Gastric Carcinoma cells)
Specification
AGS Gastric Carcinoma cells
Fixative
Formaldehyde
Permeabilization
Yes - 0.025% Triton-X in TBS
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C

Abcam user community

Verified customer

Submitted May 19 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Stomach)
Specification
Stomach
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: 10mM citrate pH6.0
Permeabilization
No
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C

Abcam user community

Verified customer

Submitted Mar 24 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Human Tissue sections (Stomach)
Specification
Stomach
Fixative
Paraformaldehyde
Permeabilization
No
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C

Abcam user community

Verified customer

Submitted Mar 18 2010

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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