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

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
IF Use a concentration of 5 µg/ml.
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 at an assay dependent concentration.
IHC-Fr 1/500.

Target

Anti-GAPDH antibody [6C5] images

  • 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 AlexaFluor®488 Goat anti-Mouse secondary (ab150117) at 2 μg/ml (shown in green) and AlexaFluor®594 Goat anti-Rabbit secondary (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:
  • Toret CP  et al. A genome-wide screen identifies conserved protein hubs required for cadherin-mediated cell-cell adhesion. J Cell Biol 204:265-79 (2014). WB ; Fruit fly (Drosophila melanogaster) . Read more (PubMed: 24446484) »
  • Zhao L  et al. Notch signaling regulates cardiomyocyte proliferation during zebrafish heart regeneration. Proc Natl Acad Sci U S A 111:1403-8 (2014). Zebrafish . Read more (PubMed: 24474765) »

See all 257 Publications for this product

Product Wall

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

Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (15% Gel)
Sample Human Tissue lysate - whole (Vastus lateralis muscle)
Specification Vastus lateralis muscle
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 18 2014

Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (12% gel)
Sample Mouse Tissue lysate - whole (Skeletal muscle)
Specification Skeletal muscle
Blocking step Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
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Submitted Jan 09 2014

Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (4-12%)
Sample Human Cell lysate - whole cell (HCT116)
Specification HCT116
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
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Submitted Dec 20 2013

Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (10)
Sample Dog Cell lysate - whole cell (MDCK)
Specification MDCK
Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
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Submitted Oct 28 2013

Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (10)
Sample Syrian Hamster Cell lysate - whole cell (BHK)
Specification BHK
Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
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Submitted Oct 28 2013

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