Overview

  • Product nameAnti-GAPDH antibody [6C5]
    See all GAPDH primary antibodies
  • Description
    Mouse monoclonal [6C5] to GAPDH
  • SpecificityThis antibody can be used as a loading control antibody, however we also recommend ab9484 as an alternative. GAPDH is a 146 kDa tetramer composed of four 30-40 kDa subunits. There is no cross-reaction with GAPDH from yeast. Preliminary data indicates that the antibody recognizes the monomer (36 kDa) and also the dimer forms of GAPDH, but not the tetrameric form of the protein.
  • Tested applicationsELISA, ICC, WB, ICC/IF, IHC-Frmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Rabbit, Chicken, Hamster, Cat, Dog, Human, Pig, Xenopus laevis, Fruit fly (Drosophila melanogaster), Fish, Monkey, Zebrafish, Baboon, African Green Monkey
    Predicted to work with: Horse, Guinea pig, Xenopus tropicalis

    Does not react with

    Goat, Cow, Saccharomyces cerevisiae
  • Immunogen

    Rabbit muscle GAPDH.

  • Positive control
    • ICC/IF: HeLa cells, NIH3T3 cells, SV40 cells

Properties

Applications

Our Abpromise guarantee covers the use of ab8245 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
ELISA Use at an assay dependent concentration.
ICC Use at an assay dependent concentration.
WB 1/500 - 1/10000. Detects a band of approximately 36 kDa (predicted molecular weight: 40.2 kDa). PubMed: 16450009
ICC/IF Use a concentration of 1 - 5 µg/ml.
IHC-Fr 1/500.

Target

Anti-GAPDH antibody [6C5] images

  • ab8245 staining GAPDH in SV40 cells. The cells were fixed with 4% formaldehyde (10min), permeabilized with 0.1% Triton X-100 for 5 minutes 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 ab8245 at 5μg/ml and ab202272 at 1/250 overnight at +4°C, followed by a further incubation at room temperature for 1h with an anti-Mouse AlexaFluor® 488 (ab150117) (shown in green). Nuclear DNA was labelled in blue with DAPI.
    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • ab8245 staining GAPDH in NIH3T3 cells. The cells were fixed with 4% formaldehyde (10min) 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 ab8245 at 1μg/ml and ab202272 at 1/250 overnight at +4°C, followed by a further incubation at room temperature for 1h with an anti-Mouse AlexaFluor® 488 (ab150117) (shown in green). Nuclear DNA was labelled in blue with DAPI.
    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • ab8245 staining GAPDH in HeLa 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 ab8245 at 5μg/ml and ab6046 at 1µg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with an anti-Mouse AlexaFluor® 488 (ab150117) at 2 μg/ml (shown in green) and anti-Rabbit AlexaFluor® 594 (ab150088) 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.

  • All lanes : Anti-GAPDH antibody [6C5] (ab8245) at 20000

    Lane 1 : Mouse hippocampus whole cell lysate
    Lane 2 : Rat hippocampus whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    HRP-conjugated Rabbit anti-mouse at 1/5000 dilution
    developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 40.2 kDa
    Observed band size : 36 kDa (why is the actual band size different from the predicted?)


    Exposure time : 10 seconds

    This image is courtesy of an anonymous Abreview

    See Abreview

  • All lanes : Anti-GAPDH antibody [6C5] (ab8245) at 2.5 µg/ml

    Lane 1 : HeLa Nuclear
    Lane 2 : HeLa whole cell lysate
    Lane 3 : A431 cell lysate
    Lane 4 : Jurkat cell lysate
    Lane 5 : HEK293 cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Alexa Fluor anti-mouse at 1/5000 dilution

    Performed under reducing conditions.

    Predicted band size : 40.2 kDa
    Observed band size : 37 kDa (why is the actual band size different from the predicted?)
    Fluorescence detection of secondary antibody.
  • All lanes : Anti-GAPDH antibody [6C5] (ab8245) at 1/20000 dilution

    Lane 1 : Rabbit aorta, whole tissue lysate at 10 µg
    Lane 2 : Blank
    Lane 3 : Mouse aorta, whole tissue lysate at 5 µg
    Lane 4 : Mouse aorta, whole tissue lysate at 7.5 µg
    Lane 5 : Mouse aorta, whole tissue lysate at 10 µg
    Lane 6 : Mouse aorta, whole tissue lysate at 12.5 µg
    Lane 7 : Mouse aorta, whole tissue lysate at 15 µg

    Secondary
    HRP conjugated sheep anti-mouse antibody at 1/10000 dilution
    developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 40.2 kDa


    Exposure time : 30 seconds

    This image is courtesy of an anonymous Abreview

    See Abreview



  • Predicted band size : 40.2 kDa

    20 µg of Raji whole cell lysate (ab7908) probed with mouse monoclonal to GAPDH (ab8245) at 10µg/ml.

    20 µg of Raji whole cell lysate (ab7908) probed with mouse monoclonal to GAPDH (ab8245) at 10µg/ml.

References for Anti-GAPDH antibody [6C5] (ab8245)

This product has been referenced in:
  • Wakuda T  et al. Perinatal asphyxia alters neuregulin-1 and COMT gene expression in the medial prefrontal cortex in rats. Prog Neuropsychopharmacol Biol Psychiatry 56:149-54 (2015). Read more (PubMed: 25194460) »
  • Durk MR  et al. Vitamin D receptor activation induces P-glycoprotein and increases brain efflux of quinidine: an intracerebral microdialysis study in conscious rats. Pharm Res 32:1128-40 (2015). Read more (PubMed: 25319098) »

See all 327 Publications for this product

Product Wall

Application Immunocytochemistry/ Immunofluorescence
Sample Rat Cell (rat neonatal myocytes)
Permeabilization Yes - 1% TRITON-X-100
Specification rat neonatal myocytes
Blocking step Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: RT°C
Fixative Paraformaldehyde
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Submitted Jun 05 2015

Application Western blot
Loading amount 25 µg
Gel Running Conditions Reduced Denaturing
Sample Human Cell lysate - whole cell (Hela, Hep G2)
Specification Hela, Hep G2
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
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Submitted Feb 09 2015

Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing
Sample Mouse Cell lysate - whole cell (liver, hepatocytes)
Specification liver, hepatocytes
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
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Submitted Feb 09 2015

Application Western blot
Loading amount 30 µg
Gel Running Conditions Reduced Denaturing
Sample Chinese Hamster Cell lysate - whole cell (CHO cell)
Specification CHO cell
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
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Submitted Oct 22 2014

Application Western blot
Loading amount 80 µg
Gel Running Conditions Reduced Denaturing (12.5% Acrylamide, 2D gel ph 3-10)
Sample Pig Tissue lysate - other (Muscle Sarcoplasm)
Specification Muscle Sarcoplasm
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
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Submitted Oct 06 2014

Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing (12% gel)
Sample Rat Tissue lysate - other (Cytosolic fractions from skeletal muscles)
Specification Cytosolic fractions from skeletal muscles
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
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Submitted Jun 02 2014

Application Western blot
Loading amount 40 µg
Gel Running Conditions Reduced Denaturing
Sample Human Cell lysate - whole cell (Colon cell line)
Specification Colon cell line
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
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Submitted May 23 2014

Application Western blot
Loading amount 25 µg
Gel Running Conditions Reduced Denaturing (4-12%)
Sample Human Cell lysate - whole cell (Colon cancer)
Specification Colon cancer
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
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Submitted Apr 14 2014

Application ELISA
Sample Human Recombinant protein (HEK293)
Specification HEK293
Type Direct
Blocking step Superblock in PBS (Pierce Cat#37580) as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10µg/mL · Temperature: 20°C
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Submitted Mar 17 2014

Application Western blot
Loading amount 10000 cells
Gel Running Conditions Reduced Denaturing (6%)
Sample Spodoptera frugiperda (SF9 cells) Cell lysate - whole cell (293 cells and Sf9 cells)
Specification 293 cells and Sf9 cells
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C
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Submitted Feb 18 2014

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