Anti-GAPDH antibody [6C5] - Loading Control (ab8245)
- Datasheet
- References (1617)
- Protocols
Overview
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Product nameAnti-GAPDH antibody [6C5] - Loading Control
See all GAPDH primary antibodies -
DescriptionMouse monoclonal [6C5] to GAPDH - Loading Control
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Host speciesMouse
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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.
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Tested applicationsSuitable for: ELISA, ICC, WB, ICC/IF, IHC-Frmore details
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Species reactivityReacts 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 tropicalis
Does not react with:
Goat, Cow, Saccharomyces cerevisiae -
Immunogen
Rabbit muscle GAPDH.
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Positive control
- ICC/IF: HeLa cells, NIH3T3 cells, SV40LT-SMC cells
Properties
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FormLiquid
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Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
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Storage bufferpH: 7.40
Preservative: 0.09% Sodium azide
Constituent: PBS -
Concentration information loading... -
PurityProtein A purified
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Purification notesChromatography on protein A Sepharose
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ClonalityMonoclonal
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Clone number6C5
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MyelomaSp2/0
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IsotypeIgG1
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Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Positive Controls
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Recombinant Protein
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Related Products
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- Anti-SDHA antibody [2E3GC12FB2AE2] (ab14715)
- GAPDH ELISA Kit (ab176642)
- Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291)
- Anti-beta Actin antibody [mAbcam 8224] - Loading Control (ab8224)
- Anti-GAPDH antibody [mAbcam 9484] - Loading Control (ab9484)
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
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FunctionHas both glyceraldehyde-3-phosphate dehydrogenase and nitrosylase activities, thereby playing a role in glycolysis and nuclear functions, respectively. Participates in nuclear events including transcription, RNA transport, DNA replication and apoptosis. Nuclear functions are probably due to the nitrosylase activity that mediates cysteine S-nitrosylation of nuclear target proteins such as SIRT1, HDAC2 and PRKDC (By similarity). Glyceraldehyde-3-phosphate dehydrogenase is a key enzyme in glycolysis that catalyzes the first step of the pathway by converting D-glyceraldehyde 3-phosphate (G3P) into 3-phospho-D-glyceroyl phosphate.
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PathwayCarbohydrate degradation; glycolysis; pyruvate from D-glyceraldehyde 3-phosphate: step 1/5.
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Sequence similaritiesBelongs to the glyceraldehyde-3-phosphate dehydrogenase family.
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Post-translational
modificationsS-nitrosylation of Cys-152 leads to interaction with SIAH1, followed by translocation to the nucleus.
ISGylated. -
Cellular localizationCytoplasm > cytosol. Nucleus. Cytoplasm > perinuclear region. Membrane. Translocates to the nucleus following S-nitrosylation and interaction with SIAH1, which contains a nuclear localization signal (By similarity). Postnuclear and Perinuclear regions.
- Information by UniProt
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Database links
- Entrez Gene: 493876 Cat
- Entrez Gene: 374193 Chicken
- Entrez Gene: 403755 Dog
- Entrez Gene: 2597 Human
- Entrez Gene: 100042025 Mouse
- Entrez Gene: 14433 Mouse
- Entrez Gene: 396823 Pig
- Entrez Gene: 100009074 Rabbit
see all -
Alternative names
- 38 kDa BFA-dependent ADP-ribosylation substrate antibody
- aging associated gene 9 protein antibody
- Aging-associated gene 9 protein antibody
see all
Images
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![Western blot - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)](https://www.abcam.com/ps/products/8/ab8245/Images/ab8245-14003-anti-gapdh-antibody-6c5-loading-control-western-blot.jpg)
Western blot - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)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. -
![Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)](https://www.abcam.com/ps/products/8/ab8245/Images/ab8245-249031-anti-gapdh-antibody-6c5-loading-control-immunofluorescence.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)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).
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![Western blot - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)](https://www.abcam.com/ps/products/8/ab8245/Images/ab8245-209020-anti-gapdh-antibody-6c5-loading-control-western-blot.jpg)
Western blot - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)This image is courtesy of an anonymous AbreviewAll 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
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![Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)](https://www.abcam.com/ps/products/8/ab8245/Images/ab8245-249030-anti-gapdh-antibody-6c5-loading-control-immunofluorescence.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)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).
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![Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)](https://www.abcam.com/ps/products/8/ab8245/Images/ab8245-228734-anti-gapdh-antibody-6c5-loading-control-immunofluorescence.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)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.
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![Western blot - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)](https://www.abcam.com/ps/products/8/ab8245/Images/ab8245-14002-anti-gapdh-antibody-6c5-loading-control-western-blot.jpg)
Western blot - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)This image is courtesy of an anonymous AbreviewAll 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
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![Western blot - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)](https://www.abcam.com/ps/products/8/ab8245/Images/ab8245_1.gif)
Western blot - Anti-GAPDH antibody [6C5] - Loading Control (ab8245)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
Protocols
References
This product has been referenced in:
- Zhao W et al. Long noncoding RNA NSCLCAT1 increases non-small cell lung cancer cell invasion and migration through the Hippo signaling pathway by interacting with CDH1. FASEB J 33:1151-1166 (2019). Read more (PubMed: 30148675) »
- Yuan J et al. GLIS2 redundancy causes chemoresistance and poor prognosis of gastric cancer based on co-expression network analysis. Oncol Rep 41:191-201 (2019). Read more (PubMed: 30320360) »
