GAPDH Antibody

Images

 
Western Blot: GAPDH Antibody [NB300-325] - Detection of mouse and rat GAPDH by western blot. Samples: Whole cell lysate (50 ug) from NIH 3T3, TCMK-1, 4T1, CT26.WT, and rat C6 cells. Antibodies: Affinity purified rabbit ...read more
Immunocytochemistry/ Immunofluorescence: GAPDH Antibody [NB300-325] - GAPDH antibody was tested in Hela cells with DyLight 488 (green). Nuclei and alpha-tubulin were counterstained with DAPI (blue) and Dylight 550 (red).
Western Blot: GAPDH Antibody [NB300-325] - Whole cell lysate (50 ug) from HeLa (H) and 293T (50 ug) probed with GAPDH Antibody diluted at 0.04 ug/ml
Western Blot: GAPDH Antibody [NB300-325] - Detection of Mouse GAPDH Whole cell lysate (5, 15 and 50 ug) from mouse NIH3T3 cells. Another Affinity purified rabbit anti-GAPDH used at 0.04 mcg/ml. Chemiluminescence with an ...read more
Simple Western: GAPDH Antibody [NB300-325] - Simple Western lane view shows a specific band for GAPDH in 0.05 mg/ml of HeLa lysate. This experiment was performed under reducing conditions using the 12-230 kDa ...read more

Product Details

Summary
Reactivity Hu, Mu, RtSpecies Glossary
Applications WB, Simple Western, ICC/IF, IP
Clonality
Polyclonal
Host
Rabbit
Conjugate
Unconjugated
Concentration
0.2 mg/ml

Order Details

GAPDH Antibody Summary

Immunogen
This GAPDH antibody was developed against an epitope between residue 300 and the C-terminus of the human GAPDH protein [accession number NP_002037.2]
Marker
Cytosolic Marker
Isotype
IgG
Clonality
Polyclonal
Host
Rabbit
Gene
GAPDH
Purity
Immunogen affinity purified
Innovator's Reward
Test in a species/application not listed above to receive a full credit towards a future purchase.

Applications/Dilutions

Dilutions
  • Immunocytochemistry/Immunofluorescence 1:250
  • Immunoprecipitation
  • Simple Western 1:500
  • Western Blot 1:2000-1:10000
Application Notes
This GAPDH antibody is useful for Immunocytochemistry/Immunofluorescence and Western blot, where a band is observed approx. 36 kDa.

In Simple Western only 10 - 15 uL of the recommended dilution is used per data point. Separated by Size-Wes, Sally Sue/Peggy Sue.
The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
Theoretical MW
36 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
Control
GAPDH Overexpression Lysate
Reviewed Applications
Read 2 Reviews rated 3.5
using
NB300-325 in the following applications:

Publications
Read Publications using
NB300-325 in the following applications:

  • WB
    3 publications

Reactivity Notes

Human and mouse.

Packaging, Storage & Formulations

Storage
Store at 4C. Do not freeze.
Buffer
TBS and 0.1% BSA
Preservative
0.09% Sodium Azide
Concentration
0.2 mg/ml
Purity
Immunogen affinity purified

Alternate Names for GAPDH Antibody

  • aging-associated gene 9 protein
  • EC 1.2.1
  • EC 1.2.1.12
  • EC 2.6.99.-
  • G3PD
  • G3PDH
  • GAPD
  • GAPDH
  • glyceraldehyde 3-phosphate dehydrogenase
  • glyceraldehyde-3-phosphate dehydrogenase
  • MGC88685
  • Peptidyl-cysteine S-nitrosylase GAPDH

Background

Glyceraldehyde 3-Phosphate Dehydrogenase (GAPDH) is a ubiquitous enzyme involved in glycolysis, converting glyceraldehyde-3-phosphate into 1,3 diphosphoglycerate. This constitutively expressed, homotetramer protein can be found in the nucleus and cytoplasm, and the monomer has a theoretical molecular weight of 36 kDa. Known as a housekeeping gene, GAPDH routinely serves as a control for real-time PCR (RT-PCR) and as a loading control for Western Blots (1-3). In addition to its role in glycolysis, GAPDH has multiple cellular functions including membrane trafficking, transcription activation, apoptosis, DNA repair, and has been implicated in various pathologies such as cancer and neurodegenerative diseases including Alzheimer's and Huntington's (4,5).

References

1) Barber RD, Harmer DW, Coleman RA, Clark BJ. (2005) GAPDH as a housekeeping gene: analysis of GAPDH mRNA expression in a panel of 72 human tissues. Physiol Genomics. 21(3):389-95. PMID: 15769908

2) Jia Y, Takimoto K. (2006) Mitogen-activated protein kinases control cardiac KChIP2 gene expression. Circ Res. 98(3):386-93. PMID: 16385079

3) Godsel LM, Hsieh SN, Amargo EV, Bass AE, Pascoe-McGillicuddy LT, Huen AC, Thorne ME, Gaudry CA, Park JK, Myung K, Goldman RD, Chew TL, Green KJ. (2005) Desmoplakin assembly dynamics in four dimensions: multiple phases differentially regulated by intermediate filaments and actin. J Cell Biol. 171(6):1045-59. PMID: 16365169

4) Sirover MA1. (1999) New insights into an old protein: the functional diversity of mammalian glyceraldehyde-3-phosphate dehydrogenase. Biochim Biophys Acta. 1432(2): 159-84. PMID: 10407139

5) Tristan C, Shahani N, Sedlak TW, Sawa A. (2011) The diverse functions of GAPDH: views from different subcellular compartments. Cell Signal. 23(2):317-23. PMID: 20727968

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

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Publications for GAPDH Antibody (NB300-325)(3)

Reviews for GAPDH Antibody (NB300-325) (2) 3.52

Average Rating: 3.5
(Based on 2 reviews)
We have 2 reviews tested in 2 species: Human, Mouse.

Reviews using NB300-325:
Filter by Applications
Simple Western
(1)
WB
(1)
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(1)
Mouse
(1)
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Images Ratings Applications Species Date Details
Simple Western GAPDH NB300-325
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2
reviewed by:
Jessica Hill
Simple Western Mouse 06/15/2018
View

Summary

ApplicationSimple Western
Sample TestedAdipose tissue
SpeciesMouse
LotA1

Comments

CommentsGAPDH pAb was used at a 1:10 concentration in subcutaneous adipose tissue (specifically the inguinal fat pad) which was loaded at a 1mg/mL concentration. The GAPDH displayed double peaks, which can be visualized in Graph view but not really apparent in Lane view). We believe this to be due to Ab burn out with chemiluminescence. This Ab was specific, and may work better for others at a lower protein concentration, or if secondary-HRP is diluted.
Western Blot GAPDH NB300-325
Enlarge
5
reviewed by:
Christopher England
WB Human 03/29/2016
View

Summary

ApplicationWestern Blot
Sample TestedHuman pancreatic cancer cells
SpeciesHuman
Lot1

Blocking

Blocking DetailsLiCor Odyssey PBS Blocker, 1X concentration, 2:00 hr, room temperature

Primary Anitbody

Dilution RatioGAPDH, 1:100, 12 hr, 4 degree Celcius, LiCor Odyssey PBS Buffer

Secondary Antibody

Secondary DescriptionGoat anti-Rabbit IgG 800 CW
Secondary Manufacturer Cat#LiCor 925-32211
Secondary Concentration1:10,000

Details

Detection NotesFluorescent Western blot, LiCor Odyssey, N/A, GADPH acted as the control for normalization

Comments

CommentsThis was a dual WB that uses two primary antibodies. The second primary antibody was: VEGF primary antibody, 1:500, 12 hr, 4 degree Celsius, LiCor Odyssey PBS BufferThe secondary antibody for this was goat anti-mouse 680 LT from LiCor at a concentration of 1:10,000.

Product General Protocols

Video Protocols

WB Video Protocol
ICC/IF Video Protocol

FAQs for GAPDH Antibody (NB300-325). (Showing 1 - 7 of 7 FAQs).

  1. We need a loading control antibody for WB working with HeLa cell line, will this GAPDH antibody work for that?
    • GAPDH is expressed in HeLa cell lines but whether or not it is the best option for a loading control antibody depends on the size of the protein you are interested in. GAPDH antibody is a good choice for low to mid MW targets, as GAPDH detects around 35 kDa.
  2. I want to know why GAPDH is used as a loading control antibody in WB techniques.
    • Our GAPDH antibody makes a good loading control because GAPDH is  a housekeeping gene essential to metabolism that is globally expressed in most tissue types and cell lines.
  3. I need a suitable loading control antibody that I can use in western blot of rat nerve extracts, which do you recommend?
    • This GAPDH antibody (NB300-221) is a good choice for a loading control antibody and has been cited in over 180 publications and used on rat lysates with good results.
  4. I'm looking for a loading control between 24kDa - 65kDa for use in simple western, we tried an actin antibody  we already had but it didn't work. What can you recommend?
    • Our GAPDH antibody NB300-221 has been validated in simple western and detects around 35 kDa so I think it would be a good choice for a simple western loading control antibody based on your size requirements.
  5. What secondary antibody should I use for visualization of the GAPDH antibody?
    • This GAPDH antibody is raised in mouse so you would want to use an anti-mouse secondary with the dye of your choice.
  6. Do you offer a smaller size for the GAPDH antibodies?
    • We offer sample sizes for many of our GAPDH loading control antibodies including NB300-221.
  7. I wanted to know which of the two housekeeping genes B-actin or GAPDH can be used for best results. I intend to do an experiment to determine expression of IL-6 and TNF-alpha in liver and kidney tissues.
    • For homogenized tissue, beta-actin and GAPDH are both fine.

Control Lysate(s)

Secondary Antibodies

 

Isotype Controls

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Blogs on GAPDH. Showing 1-10 of 13 blog posts - Show all blog posts.

Considerations for Quantitative Western blotting
By Jamshed Arslan, Pharm. D., PhD. Since its inception in 1979, Western blotting has undergone several developments. The use of radioactive probes was common throughout 1980s, but utilizing secondary antibodies labell...  Read full blog post.

The use of Beta Actin (AC-15) as a loading control across multiple species
Actin is a fundamental component of the cytoskeleton, where it has the ability to create and break down actin filament formation in response to various cell needs.  Actin has six highly conserved isoforms, however beta and gamma actin are the two...  Read full blog post.

Tips on choosing an ideal loading control antibody for Western Blotting
Western blotting is one of the most commonly used antibody assay techniques in cell and molecular biology research since its development over three decades ago, and is considered the gold standard for protein detection and quantification. When p...  Read full blog post.

GAPDH - A "housekeeping" gene with diverse functions in cellular homeostasis
Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a well-known housekeeping gene with functions in glycolysis. Many biologists are familiar with the gene and use GAPDH antibodies for a loading control when performing western blots. However, this ...  Read full blog post.

A New Standard in Antibody Testing - Simple Western Certified Antibodies
The Western blot is one of the most commonly used antibody assay techniques in cell and molecular biology research since its development over three decades ago, and is considered the gold standard for protein detection and quantification. The tradi...  Read full blog post.

GAPDH: More than a housekeeping gene
GAPDH is a 146kD tetramer glycolytic pathway metabolic enzyme composed of four 30-40 kDa subunits. It is responsible for reversibly phosphorylating its substrate glyceraldehyde 3-phosphate within the glycolytic pathway.  Apart from its role in gl...  Read full blog post.

GAPDH (Glyceraldehyde 3-Phosphate Dehydrogenase)
GAPDH is a 146 kD tetramer glycolytic pathway metabolic enzyme responsible for reversibly phosphorylating glyceraldehyde 3-phosphate. It may have other possible functions in transcriptional activation. GAPDH is highly expressed due to this housekeepin...  Read full blog post.

Time to Start Actin Like a Reliable 'Housekeeper'!
A growing body of data and studies using actin antibodies supports a view of the actin cytoskeleton of smooth muscle cells as a dynamic structure that plays an integral role in regulating the development of mechanical tension and the material properti...  Read full blog post.

Beta Actin and GAPDH: The Importance of Loading Controls
Western blotting is an essential technique to probe protein expression in complex cell or tissue lysates. To accurately determine protein expression and interpret Western blot results, it is important to use loading controls. A loading control anti...  Read full blog post.

The GAPDH Antibody in Western blot Assays
The loading controls on our antibody database are widely used in gel electrophoresis and Western blotting studies. Products like the GAPDH antibody detect "housekeeping" proteins which are abundantly distributed in cells. This makes them useful for ch...  Read full blog post.

Showing 1-10 of 13 blog posts - Show all blog posts.

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Recent Reviews

3.5
5
1
4
0
3
0
2
1
1
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Jessica Hill
06/15/2018
Application: Simple Western
Species: Mouse

Christopher England
03/29/2016
Application: WB
Species: Human

Bioinformatics

Gene Symbol GAPDH
Entrez
Uniprot