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Datasheet
Peptide-affinity Purified Polyclonal Antibody to GAPDH - Loading Control
Cat.No IMG-3073
Description GAPDH - Loading Control
Format Purified, Peptide Affinity
Unit 0.1 mg
Price(USD) 295.00
MSDS
Catalog No
:
IMG-3073
Contents
:
0.1 mg of purified antibody. 0.5 mg/ml in Tris saline, 0.02% sodium azide, pH7.3 with 0.5% bovine serum albumin.
Isotype
:
Goat Ig
Clone
:
N/A
Purification
:
Antigen Affinity Chromatography
Species
:
Cow, Dog, Drosophila, Human, Monkey, Mouse, Pig, Rat
Predicted React
:
Xenopus
Host
:
Goat
Application
ELISA: >1:16000 IHC (paraffin): 0.3µg/ml Western blot analysis: 0.01-0.1 ug/ml Storage Store at -20°C. Recommended Positive Control : Brain, heart, kidney, liver, lung, stomach, spleen, ovary, testis, and 293. Most cell lines and tissues will be positive for GAPDH.
Background Glyceraldehyde 3-Phosphate Dehydrogenase (GAPDH, G3PDH or GPDH) is one of the key enzymes involved in glycolysis; it catalyzes the reversible oxidative phosphorylation of glyceraldehydes-3-phosphate.
The GAPDH gene is constitutively and stably expressed at high levels in almost all tissues and cells, and as such is considered to be a “housekeeping” gene. Housekeeping proteins like GAPDH are useful as loading controls for western blots and protein normalization.
Antigen
Peptide with sequence HQVVSSDFNSDT, from C Terminus of the protein sequence according to NP_002037.
Application Notes
On Western blots, GAPDH is detected as a band of approximately 36-40 kDa. GAPDH antibodies are widely used as loading controls for quantitative Western blotting, including normalizing Western blot results to GAPDH. GAPDH is highly conserved across species, and antibodies to GAPDH typically have broad species reactivity.
IHC staining of paraffin embedded Human Tonsil using IMG-3073 at 0.3µg/ml. Microwaved antigen retrieval with Tris/EDTA buffer pH9, HRP-staining. A) Staining of germinal centre cells. B) Staining of endothelial cells.
Application of the GAPDH antibody as a protein loading control in Western blots. Total proteins from various mouse tissue lysates were normalized using the GAPDH polyclonal antibody (IMG-3073 at 0.05 ug/ml). Mouse tissue lysates, Cat nos: Brain (40101), Heart(40102), Kidney (40104), Liver (40105), Lung (40106), Stomach (40110), Spleen (40109), Ovary (40108), and Testis (40111).
Application of the GAPDH antibody as a protein loading control in Western blots. Total proteins from various human tissue lysates were normalized using the GAPDH polyclonal antibody (IMG-3073 at 0.05 ug/ml). Human tissue lysates, Cat nos: Kidney (40144), Liver (40145), Stomach (40152), and Testis (40150).
Reference 1. Burke JR, Enghild JJ, Martin ME, Jou YS, Myers RM, Roses AD, Vance JM,Strittmatter WJ. Huntingtin and DRPLA proteins selectively interact with the enzyme GAPDH. Nat Med. 1996 Mar;2(3):347-50. PMID: 8612237 2. Fortun, J., W.A. Dunn, S. Joy, J. Li, and L. Notterpek. 2003. Emerging role for autophagy in the removal of aggresomes in Schwann Cells. J. Neurosci. 23: 10672-10680. 3. Ellis,R.C., J.N. Earnhardt, R.L. Hayes, K.K. Wang, and D.K. Anderson. 2004. Cathepsin B mRNA and protein expression following contusion spinal cord injury in rats. J. Nuerochem. 88:689-697. 4. Kiepe D. et al.: Defined carboxy-terminal fragments of insulin-like growth factor (IGF) binding protein 2 exert similar mitogenic activity on cultured rat growth plate chondrocytes as IGF-I. Endocrinology. 2008 Oct;149(10):4901-11. Epub 2008 Jun 12. PMID: 18556354
Product Citations 1. Mice lacking NKCC1 are protected from development of bacteremia and hypothermic sepsis secondary to bacterial pneumonia. Nguyen M, A Pace, and B Koller The Journal of Experimental Medicine 204:1383-1393 (2007).WB, mouse lung tissue lysates (used as a protein loading control): Fig 8. 2. Brain fatty acid binding protein (Fabp7) is diurnally regulated in astrocytes and hippocampal granule cell precursors in adult rodent brain. Gerstner J, B Quentin, W Heyden, T Lavaute, J Yin, C Landry. PLoS ONE 3: e1631 (2008). Imgenex antibodies cited [WB (mouse brain tissue)]: 1. GAPDH - Loading Control (IMG-3073): Figs. 1A,B; 3. 2. b-actin (IMG-5142A): Fig. 3. 3. Developmental regulation of the NMDA receptor subunits, NR3A and NR1, in human prefrontal cortex. Henson M, A Roberts, K Salimi, S Vadlamudi, R Hamer, J Gilmore, L Jarskog, B Philpot. Cerebral Cortex doi:10.1093/cercor/bhn017 (2008). WB (mouse frontal cortex lysates), Fig. 1. 4. Distinct functions for the catalytic and hemopexin domains of a Drosophila matrix metalloproteinase. Glasheen BM, AT Kabra, A Page-McCaw. PNAS 106:2659-2664 (2009). WB: Fig 4A (drosophila embryos), Fig 4B (drosophila larvae). Note: GAPDH was used as a loading control. Differential expression of GAPDH isoforms were seen in embryos (one GAPDH band between 36 and 50 kDa) compared to larvae (two GAPDH bands between 31 and 37 kDa).
Research purposes only.
Not for diagnostic or in vivo use. This product is guaranteed to perform as indicated on the datasheet for one year from the date of purchase.
The IMGENEX Knowledgebase Center is an interactive tool containing hundreds of answers to frequently
asked technical questions compiled by IMGENEX technical support scientists. Updated daily, you can quickly
find information about specific products and applications.
IMG-3073 [Peptide-affinity Purified Polyclonal Antibody to GAPDH - Loading Control]
What is the difference between b-actin and b-tubulin Abs for normalization of proteins?
Answer :
The concept of using proteins for normalization/internal reference is based on the premise that certain proteins are expressed at high levels and that their levels remain relatively constant whereas the levels of other proteins may change. Therefore, this set of proteins, commonly referred as "housekeeping" proteins have become useful for normalization/standardization. These proteins commonly include b-Actin, b-Tubulin, a Gapdh. In some assays, researchers may select more than one protein or a cocktail of proteins to normalize against.
However, you should be aware that one caveat with using housekeeping proteins for protein normalization is like other genes in some pathological or experimental conditions, their expression may vary because they have roles in more than one cellular function. For example, for GAPDH is involved in the glycolytic pathway, and tubulin and actin in the cytoskeleton.
Studies concerning the choice of the housekeeping gene best-suited as internal reference have been carried out. I would encourage you to refer to the published literature for additional information on the the topic, eg "Housekeeping proteins: A preliminary study illustrating some limitations as useful references in protein expression studies: Roisean E. Ferguson, Helen P. Carroll 1, Adrian Harris 2, Eamonn R. Maher 3, Peter J. Selby 1, Rosamonde E. Banks 1 *Volume 5, Issue 2 , Pages 566 - 571, 2005. (attached for your convenience).
This and like publications should be able to help you determine what housekeeping protein (eg actin, tubulin, gapdh or even another protein) is commonly used for standardizatin in the model system, including cell type or tissue type that you are you are using.
The follow Imgenex antibodies have been designed to be useful for normalization of proteins:
I would encourage you to look at the data sheets for them to determine which one(s) may be most useful for you. For example, GAPDH (IMG-5019A-1/2) http://www.imgenex.com/antibody_details.php?catalogeqtoIMG-5019A-1 has several Product Citations where the researchers have use the antibody for protein normalization/standardization. Beta-actin (IMG-5142) http://www.imgenex.com/antibody_details.php?catalogeqtoIMG-5142A also has several Product Citations. Product citations are examples where researchers have purchased that particular product and published with it.
