Overview

  • Product name

    Anti-GAPDH antibody [6C5] - Loading Control
    See all GAPDH primary antibodies
  • Description

    Mouse monoclonal [6C5] to GAPDH - Loading Control
  • Host species

    Mouse
  • Specificity

    This GAPDH antibody can be used as a loading control antibody. 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 GAPDH antibody- loading control ab8245 recognizes the monomer (36 kDa) and also the dimer forms of GAPDH, but not the tetrameric form of the protein.

  • Tested applications

    Suitable for: ELISA, ICC, WB, ICC/IF, IHC-Frmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Rabbit, Chicken, Hamster, Cat, Dog, Human, Pig, Xenopus laevis, Fish, Monkey, Zebrafish, Baboon, African green monkey
    Predicted to work with: Horse, Guinea pig, Xenopus tropicalisDoes not react with: Goat, Cow, Saccharomyces cerevisiae
  • Immunogen

    Rabbit muscle GAPDH.

  • Positive control

    • ICC/IF: HeLa cells, NIH3T3 cells, SV40LT-SMC 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

Images

  • All lanes : Anti-GAPDH antibody [6C5] - Loading Control (ab8245)

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

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : 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

    See Abreview

  • ab8245 staining GAPDH in SV40LT-SMC cells.

    The cells were fixed with 4% formaldehyde (10 minutes), 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 1 hour. 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 Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed (ab150117) (shown in green). Nuclear DNA was labeled in blue with DAPI.

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • Anti-GAPDH antibody [6C5] - Loading Control (ab8245) at 10 µg/ml + Raji (Human Burkitt's lymphoma cell line) whole cell lysate at 20 µg

    Predicted band size: 40.2 kDa

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

    Lane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) Nuclear
    Lane 2 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
    Lane 3 : A431 (Human epidermoid carcinoma cell line) cell lysate
    Lane 4 : Jurkat (Human T cell leukemia cell line from peripheral blood) cell lysate
    Lane 5 : HEK-293 (Human epithelial cell line from embryonic kidney) cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : 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.
  • ab8245 staining GAPDH in NIH/3T3 (Mouse embryo fibroblast cell line) cells.

    The cells were fixed with 4% formaldehyde (10 minutes) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1 hour. The cells were then incubated with ab8245 at 1 μg/ml and ab202272 at 1/250 overnight at +4°C, followed by a further incubation at room temperature for 1 hour with Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed (ab150117) (shown in green). Nuclear DNA was labeled in blue with DAPI.

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • All lanes : Anti-GAPDH antibody [6C5] - Loading Control (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
    All lanes : 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

    See Abreview

  • ab8245 staining GAPDH in HeLa (Human epithelial cell line from cervix adenocarcinoma) cells.

    The cells were fixed with 100% methanol (5 minutes) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1 hour. The cells were then incubated with ab8245 at 5 μg/ml and ab6046 at 1 µg/ml overnight at +4°C, followed by a further incubation at room temperature for 1 hour with Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed (ab150117) at 2 μg/ml (shown in green) and Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed (ab150088) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labeled 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.

References

This product has been referenced in:

  • He B  et al. MicroRNA-326 decreases tau phosphorylation and neuron apoptosis through inhibition of the JNK signaling pathway by targeting VAV1 in Alzheimer's disease. J Cell Physiol 235:480-493 (2020). Read more (PubMed: 31385301) »
  • Sun H  et al. miR-4429 sensitized cervical cancer cells to irradiation by targeting RAD51. J Cell Physiol 235:185-193 (2020). Read more (PubMed: 31190335) »
See all 1717 Publications for this product

Customer reviews and Q&As

1-10 of 101 Abreviews or Q&A

Application
Western blot
Sample
Rat Tissue lysate - whole (Brain tissue)
Gel Running Conditions
Reduced Denaturing (8% SDS Running gel)
Loading amount
80 µg
Treatment
10mg ketamine
Specification
Brain tissue
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Collin Gagne

Verified customer

Submitted Nov 12 2019

Application
Western blot
Sample
Human Tissue lysate - whole (skin tissue)
Gel Running Conditions
Reduced Denaturing (10% gel)
Loading amount
30 µg
Treatment
UV B (100mJ/cm2)
Specification
skin tissue
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Apr 08 2019

Application
Western blot
Sample
Mouse Cell lysate - other (MEF (cytosol fraction))
Gel Running Conditions
Reduced Denaturing (12)
Loading amount
10 µg
Specification
MEF (cytosol fraction)
Blocking step
Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Jan 09 2019

Application
Western blot
Sample
Human Cell lysate - whole cell (H1299 cells)
Gel Running Conditions
Reduced Denaturing (12)
Loading amount
25 µg
Specification
H1299 cells
Blocking step
Licor Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Sep 21 2018

Application
Western blot
Sample
Mouse Tissue lysate - whole (Brain, heart, liver)
Gel Running Conditions
Reduced Denaturing (12% gel)
Loading amount
25 µg
Specification
Brain, heart, liver
Blocking step
Licor Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Sep 21 2018

Application
Western blot
Sample
Human Cell lysate - other (A549)
Gel Running Conditions
Reduced Denaturing (10%)
Loading amount
50 µg
Specification
A549
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C

Abcam user community

Verified customer

Submitted May 22 2018

Application
Western blot
Sample
Rat Tissue lysate - whole (rat hippocampus tissue)
Gel Running Conditions
Non-reduced Denaturing
Loading amount
50 µg
Treatment
RIPA buffer
Specification
rat hippocampus tissue
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Sep 14 2017

Application
Western blot
Sample
Bos taurus Tissue lysate - whole (Lane 1Bovine Liver and Lane 2 Bovine mammary gland)
Gel Running Conditions
Reduced Denaturing (4-15% Gradient)
Loading amount
20 µg
Specification
Lane 1Bovine Liver and Lane 2 Bovine mammary gland
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Julie Kim

Verified customer

Submitted May 12 2017

Application
Western blot
Sample
Mouse Tissue lysate - whole (Mouse whole brain)
Gel Running Conditions
Reduced Denaturing
Loading amount
20 µg
Specification
Mouse whole brain
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted May 04 2017

Application
Western blot
Sample
Rat Tissue lysate - whole (Liver)
Gel Running Conditions
Reduced Denaturing (12%)
Loading amount
50 µg
Specification
Liver
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Feb 24 2017

1-10 of 101 Abreviews or Q&A

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