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  • Product Name
    GAPDH Antibody
  • Catalog No.
    AF7021
  • RRID
    AB_2839421
  • Source
    Rabbit
  • Application
    WB,IHC,IF/ICC,ELISA
  • Reactivity
    Human, Mouse, Rat, Pig, Bovine, Goat, Monkey, Chicken
  • Prediction
    Horse(100%), Sheep(100%), Rabbit(100%), Dog(100%), Xenopus(90%)
  • UniProt
  • Mol.Wt
    (Observed)37kD; (Calculated)36kDa
  • Concentration
    1mg/ml
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Product Information

Alternative Names:Expand▼

GAPDH, A1 40 kd subunit, Activator 1 40 kd subunit, G3PD, GAPD, G3pdh, Rfc40, Rf-c 40 kd subunit

Applications:

WB 1:3000-1:30000, IHC 1:200, IF/ICC 1:200, ELISA(peptide) 1:20000-1:40000

Reactivity:

Human, Mouse, Rat, Pig, Bovine, Goat, Monkey, Chicken

Predicted Reactivity:

Horse(100%), Sheep(100%), Rabbit(100%), Dog(100%), Xenopus(90%)

Source:

Rabbit

Clonality:

Polyclonal

Purification:

The antiserum was purified by peptide affinity chromatography using SulfoLink™ Coupling Resin (Thermo Fisher Scientific).

Specificity:

GAPDH antibody detects endogenous levels of total GAPDH.

RRID:

AB_2839421
Please cite this product as: Affinity Biosciences Cat# AF7021, RRID:AB_2839421.

Format:

Liquid

Concentration:

1mg/ml

Storage Condition and Buffer:

Rabbit IgG in phosphate buffered saline , pH 7.4, 150mM NaCl, 0.02% sodium azide and 50% glycerol.Store at -20 °C.Stable for 12 months from date of receipt.

Immunogen Information

Immunogen:

A synthesized peptide derived from human GAPDH.

Uniprot:



>>Visit The Human Protein Atlas

Gene ID:

Gene Name:

GAPDH

Molecular Weight:

Observed Mol.Wt.: (Observed)37kD.
Predicted Mol.Wt.: (Calculated)36kDa.

Subcellular Location:

Cytoplasm > 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.

Description:

Glyceraldehyde 3 phosphate dehydrogenase (GAPDH) iswell known as one of the key enzymes involved in glycolysis.As well as functioning as a glycolytic enzyme in cytoplasm,recent evidence suggests that mammalian GAPDHis also involved in a great number of intracellular procesessuch as membrane fusion, microtubule bundling, phosphotransferaseactivity, nuclear RNA export, DNA replication,and DNA repair.

Sequence:
MGKVKVGVNGFGRIGRLVTRAAFNSGKVDIVAINDPFIDLNYMVYMFQYDSTHGKFHGTVKAENGKLVINGNPITIFQERDPSKIKWGDAGAEYVVESTGVFTTMEKAGAHLQGGAKRVIISAPSADAPMFVMGVNHEKYDNSLKIISNASCTTNCLAPLAKVIHDNFGIVEGLMTTVHAITATQKTVDGPSGKLWRDGRGALQNIIPASTGAAKAVGKVIPELNGKLTGMAFRVPTANVSVVDLTCRLEKPAKYDDIKKVVKQASEGPLKGILGYTEHQVVSSDFNSDTHSSTFDAGAGIALNDHFVKLISWYDNEFGYSNRVVDLMAHMASKE

Research Background

Function:

Has 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. Modulates the organization and assembly of the cytoskeleton. Facilitates the CHP1-dependent microtubule and membrane associations through its ability to stimulate the binding of CHP1 to microtubules (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. Component of the GAIT (gamma interferon-activated inhibitor of translation) complex which mediates interferon-gamma-induced transcript-selective translation inhibition in inflammation processes. Upon interferon-gamma treatment assembles into the GAIT complex which binds to stem loop-containing GAIT elements in the 3'-UTR of diverse inflammatory mRNAs (such as ceruplasmin) and suppresses their translation.

Post-translational Modifications:

S-nitrosylation of Cys-152 leads to interaction with SIAH1, followed by translocation to the nucleus (By similarity). S-nitrosylation of Cys-247 is induced by interferon-gamma and LDL(ox) implicating the iNOS-S100A8/9 transnitrosylase complex and seems to prevent interaction with phosphorylated RPL13A and to interfere with GAIT complex activity.

ISGylated.

Sulfhydration at Cys-152 increases catalytic activity.

Oxidative stress can promote the formation of high molecular weight disulfide-linked GAPDH aggregates, through a process called nucleocytoplasmic coagulation. Such aggregates can be observed in vivo in the affected tissues of patients with Alzheimer disease or alcoholic liver cirrhosis, or in cell cultures during necrosis. Oxidation at Met-46 may play a pivotal role in the formation of these insoluble structures. This modification has been detected in vitro following treatment with free radical donor (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide. It has been proposed to destabilize nearby residues, increasing the likelihood of secondary oxidative damages, including oxidation of Tyr-45 and Met-105. This cascade of oxidations may augment GAPDH misfolding, leading to intermolecular disulfide cross-linking and aggregation.

Succination of Cys-152 and Cys-247 by the Krebs cycle intermediate fumarate, which leads to S-(2-succinyl)cysteine residues, inhibits glyceraldehyde-3-phosphate dehydrogenase activity. Fumarate concentration as well as succination of cysteine residues in GAPDH is significantly increased in muscle of diabetic mammals. It was proposed that the S-(2-succinyl)cysteine chemical modification may be a useful biomarker of mitochondrial and oxidative stress in diabetes and that succination of GAPDH and other thiol proteins by fumarate may contribute to the metabolic changes underlying the development of diabetes complications.

Subcellular Location:

Cytoplasm>Cytosol. Nucleus. Cytoplasm>Perinuclear region. Membrane. Cytoplasm>Cytoskeleton.
Note: Translocates to the nucleus following S-nitrosylation and interaction with SIAH1, which contains a nuclear localization signal (By similarity). Postnuclear and Perinuclear regions.

Extracellular region or secreted Cytosol Plasma membrane Cytoskeleton Lysosome Endosome Peroxisome ER Golgi apparatus Nucleus Mitochondrion Manual annotation Automatic computational assertionGraphics by Christian Stolte

Subunit Structure:

Homotetramer. Interacts with TPPP; the interaction is direct. Interacts (when S-nitrosylated) with SIAH1; leading to nuclear translocation. Interacts with RILPL1/GOSPEL, leading to prevent the interaction between GAPDH and SIAH1 and prevent nuclear translocation. Interacts with CHP1; the interaction increases the binding of CHP1 with microtubules. Associates with microtubules (By similarity). Interacts with EIF1AD, USP25, PRKCI and WARS1. Interacts with phosphorylated RPL13A; inhibited by oxidatively-modified low-densitity lipoprotein (LDL(ox)). Component of the GAIT complex. Interacts with FKBP6; leading to inhibit GAPDH catalytic activity.

Similarity:

The [IL]-x-C-x-x-[DE] motif is a proposed target motif for cysteine S-nitrosylation mediated by the iNOS-S100A8/A9 transnitrosylase complex.

Belongs to the glyceraldehyde-3-phosphate dehydrogenase family.

Research Fields

Research Fields:

· Environmental Information Processing > Signal transduction > HIF-1 signaling pathway.(View pathway)
· Human Diseases > Neurodegenerative diseases > Alzheimer's disease.
· Metabolism > Global and overview maps > Biosynthesis of amino acids.
· Metabolism > Global and overview maps > Carbon metabolism.
· Metabolism > Carbohydrate metabolism > Glycolysis / Gluconeogenesis.
· Metabolism > Global and overview maps > Metabolic pathways.

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Application: WB    Species:human;    Sample:Not available

Figure 5. LXA4 pretreatment downregulates the LPS-induced secretion and expression of HMGB1 in keratinocytes. NHEKs were stimulated with LPS (10μg/ml) with and without preincubation with LXA4 (100nmol/l) for 30minutes. (A) Western blot analysis shows specifc bands for the expression of nuclear and cytoplasmic HMGB1 in NHEKs. Full-length blots are presented in Suppl.


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Application: WB    Species:Not available;    Sample:Not available


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Application: WB    Species:human;    Sample:Not available

FIGURE 4. Knockdown of HIF-1α interferes with hypoxia-induced autophagy in HESCs. (A) Representative western blots of HIF-1α, Beclin1 and LC3 protein in HESCs transfected with scrambled control siRNA or HIF- 1α specific siRNA in the presence or absence of hypoxia. (B) The protein expression levels were quantified by Image J software and normalized to GAPDH protein levels. The data are presented as the means ± SD from at least three independent experiments (*p<0.05;**p<0.01; ***p


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Application: WB    Species:human;    Sample:SW480


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Application: WB    Species:human;    Sample:human

Fig. 2. Transfection and reagents influenced the expression of ephrin-B2 in EPCs.


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90). Su C et al. [EXPRESS] X-ray induces mechanical and heat allodynia in mouse via TRPA1 and TRPV1 activation. Mol Pain 2019 Apr 23:1744806919849201 (PubMed: 31012378)

91). Zhou Y et al. Magnesium ion leachables induce a conversion of contractile vascular smooth muscle cells to an inflammatory phenotype. J Biomed Mater Res B Appl Biomater 2018 Oct 1 (PubMed: 30270501)

92). Chen D et al. Quxie Capsule Inhibits Colon Tumor Growth Partially Through Foxo1-Mediated Apoptosis and Immune Modulation. Integr Cancer Ther 2019 Jan-Dec;18:1534735419846377 (PubMed: 31030593)

93). Bai XD et al. Constructing a better binding peptide for drug delivery targeting the interleukin-4 receptor. J Drug Target 2020 May 4:1-26 (PubMed: 32363946)

94). Ding L et al. The protective effects of polysaccharide extract from Xin-Ji-Er-Kang formula on Ang II-induced HUVECs injury, L-NAME-induced hypertension and cardiovascular remodeling in mice. BMC Complement Altern Med 2019 Jun 13;19(1):127 (PubMed: 31196042)

95). Zhao X et al. Up-regulation of miR-21 and miR-23a Contributes to As2 O3 -induced hERG Channel Deficiency. Basic Clin Pharmacol Toxicol 2015 Jun;116(6):516-23 (PubMed: 25395240)

Application: WB    Species:Not available;    Sample:Not available


96). Ruan H et al. Anlotinib attenuated bleomycin-induced pulmonary fibrosis via the TGF-β1 signalling pathway. J Pharm Pharmacol 2019 Oct 28 (PubMed: 31659758)

97). Wang L et al. CDK3 is a major target of miR-150 in cell proliferation and anti-cancer effect. Exp Mol Pathol 2017 Apr;102(2):181-190 (PubMed: 28108217)

Application: WB    Species:human;    Sample:H1299 cells

Figure 6. miR-150 suppresses CDK3 expression in lung cancer compared cells. SPC1and H1299 cells were transfected with miR-150 mimics and mimics NC for forty-eight hours, respectively. A. Western blot of CDK3 in SPC1and H1299 cells. B. qRT-PCR analysis of miR-150 in SPC1and H1299 cells. C. qRT-PCR analysis of CDK3 mRNA in SPC1and H1299 cells. Data are the means ± SEM of three independent experiments. *p<0.05, **p


98). Cheng Z et al. Rab1A promotes proliferation and migration abilities via regulation of the HER2/AKT-independent mTOR/S6K1 pathway in colorectal cancer. Oncol Rep 2019 Mar 15 (PubMed: 30896866)

99). Gao T et al. Aberrant hypomethylation and overexpression of the eyes absent homologue 2 suppresses tumor cell growth of human lung adenocarcinoma cells. Oncol Rep 2015 Nov;34(5):2333-42 (PubMed: 26329363)

Application: WB    Species:human;    Sample:NSCLC cells

Figure 1. Expression of EYA2 in NSCLC cells. (A) Western blot analysis. (B) EYA2 protein expression was significantly upregulated 3.2‑fold in the A549 cells compared to the BEAS-2B cells, but had no obvious change in the H661 and SKMES1 cells. (C) mRNA level of EYA2 exhibited a 2.1-fold upregulation in the A549 cells compared to the BEAS-2B cells. The groups were obtained from different parts of the same gel. * P


100). Zhao H et al. KIF15 promotes bladder cancer proliferation via the MEK-ERK signaling pathway. Cancer Manag Res 2019 Feb 26;11:1857-1868 (PubMed: 30881113)

101). Liu Y et al. Endovascular hypothermia improves post-resuscitation myocardial dysfunction by increasing mitochondrial biogenesis in a pig model of cardiac arrest. Cryobiology 2019 Jul 6 (PubMed: 31283936)

102). Zhao WX et al. Notch2 and Notch3 suppress the proliferation and mediate invasion of trophoblast cell lines. Biol Open 2017 Aug 15;6(8):1123-1129 (PubMed: 28606936)

103). Zhang J et al. Long non-coding RNA NEAT1 regulates E2F3 expression by competitively binding to miR-377 in non-small cell lung cancer. Oncol Lett 2017 Oct;14(4):4983-4988 (PubMed: 29085511)

104). Wu Y et al. Study on the mechanism of JAK2/STAT3 signaling pathway-mediated inflammatory reaction after cerebral ischemia. Mol Med Rep 2018 Apr;17(4):5007-5012 (PubMed: 29393445)

105). Mao X et al. Inhibitors of PARP-1 exert inhibitory effects on the biological characteristics of hepatocellular carcinoma cells in vitro. Mol Med Rep 2017 Jul;16(1):208-214 (PubMed: 28498459)

Application: WB    Species:human;    Sample:HepG2

Figure 4. Effects of different concentrations of AG014699 and BSI‑201 on protein levels of Caspase 3, Caspase 8, Bax and Bcl‑2 in HepG2 cells.(A) Blots showing proteins in cells treated with AGO14699 and (B) quantification. (C) Blots showing proteins in cells treated with (C) BSI‑201 and (D) quantification. * P<0.05, compared with the control group; ∆P<0.05, compared with the low dose group; ∆∆P<0.05, compared with the middle dose group. CTRL, control; Bcl‑2, B‑cell lymphoma 2; BAX, Bcl‑2‑associated X protein.


106). Xu Y et al. Novel conjugates of endoperoxide and 4-anilinoquinazoline induce myeloma cell apoptosis by inhibiting the IGF1-R/AKT/mTOR signaling pathway. Biosci Trends 2020 Mar 13 (PubMed: 32173687)

107). Jiang W et al. Identification of Key Candidate Genes and Pathways of Candida albicans-Infected Human Umbilical Vein Endothelial Cells and Drug Screening. Indian J Microbiol 2020 Mar;60(1):62-69 (PubMed: 32089575)

108). Ye R et al. Stress causes cognitive impairment by affecting cholesterol efflux and reuptake leading to abnormalities in lipid metabolism of rats. J Integr Neurosci 2020 Mar 30;19(1):39-49 (PubMed: 32259885)

109). Liu S et al. Isosteroid alkaloids from Fritillaria cirrhosa bulbus as inhibitors of cigarette smoke-induced oxidative stress. Fitoterapia 2020 Jan;140:104434 (PubMed: 31760067)

110). Liu YR et al. Twist1 confers multidrug resistance in colon cancer through upregulation of ATP-binding cassette transporters. Oncotarget 2017 May 2;8(32):52901-52912 (PubMed: 28881781)

111). Luo ML et al. Conditioned Medium from Human Umbilical Vein Endothelial Cells Promotes Proliferation, Migration, Invasion and Angiogenesis of Adipose Derived Stem Cells. Curr Med Sci 2018 Feb;38(1):124-130 (PubMed: 30074161)

112). Li XH et al. Parthenolide attenuated bleomycin-induced pulmonary fibrosis via the NF-κB/Snail signaling pathway. Respir Res 2018 Jun 5;19(1):111 (PubMed: 29871641)

113). Shen B et al. Aucubin inhibited lipid accumulation and oxidative stress via Nrf2/HO-1 and AMPK signalling pathways. J Cell Mol Med 2019 Apr 4 (PubMed: 30950217)

114). Shen B et al. Geniposide alleviates non-alcohol fatty liver disease via regulating Nrf2/AMPK/mTOR signalling pathways. J Cell Mol Med 2020 Apr 15 (PubMed: 32293113)

115). Sun C et al. Hsa-miR-326 targets CCND1 and inhibits non-small cell lung cancer development. Oncotarget 2016 Feb 16;7(7):8341-59 (PubMed: 26840018)

Application: WB    Species:Not available;    Sample:Not available

Figure 8: Ectopic expression of miR-326 promotes apoptosis in A549 and SPC-A-1 cells. (C) Western-blot of Bcl2 protein in A549 and SPC-A-1 cells after transfection.


116). Xie J et al. circEPSTI1 regulates ovarian cancer progression via decoying miR-942. J Cell Mol Med 2019 Mar 19 (PubMed: 30887698)

117). Xin XL et al. Mechanisms of IFNalpha-1a-Induced Apoptosis in a Laryngeal Cancer Cell Line. Med Sci Monit 2019 Sep 22;25:7100-7114 (PubMed: 31542790)

118). Xiang XJ et al. MiR-1271 Inhibits Cell Proliferation, Invasion and EMT in Gastric Cancer by Targeting FOXQ1. Cell Physiol Biochem 2015;36(4):1382-94 (PubMed: 26159618)

Application: WB    Species:human;    Sample:human gastric cancer

Fig. 2. FOXQ1 is a direct target of miR-1271. (A) TargetScan (www.targetscan. org) and miRanda (www. microRNA.org) showed that FOXQ1 was a direct target of miR-1271. (B) The luciferase reporter assay showed that FOXQ1 could be targeted by miR-1271 in MGC- 803 and SGC-7901 cells. (C) The qPCR and western blot analyses showed that miR-1271 mimics could inhibit the mRNA and protein expression of FOXQ1 in MGC-803 cells, whereas the miR-1271 inhibitor could increase the mRNA and protein expression of FOXQ1 in SGC-7901 cells. Three independent experiments were conducted. *P < 0.05, **P < 0.01.


119). Liu H et al. Long non-coding RNA MALAT1 mediates hypoxia-induced pro-survival autophagy of endometrial stromal cells in endometriosis. J Cell Mol Med 2019 Jan;23(1):439-452 (PubMed: 30324652)

120). et al. Autophagy, lysosome dysfunction and mTOR inhibition in MNU-induced photoreceptor cell damage.

121). et al. Prediction of crucial epigenetically‑associated, differentially expressed genes by integrated bioinformatics analysis and the identification of S100A9 as a novel biomarker in psoriasis.

122). et al. Evaluation of curcumin-mediated photodynamic therapy on the reverse of multidrug resistance in tumor cells.

123). Wang H;Zhang S;Yang F;Xin R;Wang S;Cui D;Sun Y; et al. The gut microbiota confers protection in the CNS against neurodegeneration induced by manganism. Biomed Pharmacother 2020 Apr 21;127:110150. (PubMed: 32330797)

124). et al. Arsenic induces pancreatic dysfunction and ferroptosis via mitochondrial ROS-autophagy-lysosomal pathway.

125). Xiulong Niu et al. IL-17A exacerbates cisplatin-based resistance of OVCA via upregulating the expression of ABCG2 and MDR1 through Gli1-mediated Hh signaling. Oncotarget 2016

126). et al. Effects of Agaricus blazei Murrill polysaccharides on hyperlipidemic rats by regulation of intestinal microflora.

127). Sun H et al. Role of 5-HT1A receptor in insular cortex mediating stress - induced visceral sensory dysfunction. Neurogastroenterol Motil 2016 Jul;28(7):1104-13 (PubMed: 26969829)

Application: WB    Species:Not available;    Sample:Not available

Figure 3 Comparison of the expression levels of HTR1As and NR2B in bilateral insular cortexes among three groups. The mRNA and protein expression levels of HTR1As (A and C) and NR2B (B and D) in bilateral insular cortexes were compared among WAS, sham WAS and normal control groups. (E) and (F): Representative immunoblots from the bilateral insular cortexes show the expression of NR2B, 5HTR1A and GAPDH, with an indication of size (kilodaltons, kDa). *represented that the difference was significant, p < 0.05. WAS, water avoidance stress.


128). Zhao L;Bi M;Zhang H;Shi M; et al. Downregulation of NEAT1 Suppresses Cell Proliferation, Migration, and Invasion in NSCLC Via Sponging miR-153-3p. Cancer Biother Radiopharm 2020 May 7. (PubMed: 32380843)

129). et al. Cdk5-Mediated Drp1 Phosphorylation Drives Mitochondrial Defects and Neuronal Apoptosis in Radiation-Induced Optic Neuropathy.

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Catalog Number :

AF7021-BP
(Blocking peptide available as AF7021-BP)

Price/Size :

$350/1mg.
Tips: For phospho antibody, we provide phospho peptide(0.5mg) and non-phospho peptide(0.5mg).

Function :

Blocking peptides are peptides that bind specifically to the target antibody and block antibody binding. These peptide usually contains the epitope recognized by the antibody. Antibodies bound to the blocking peptide no longer bind to the epitope on the target protein. This mechanism is useful when non-specific binding is an issue, for example, in Western blotting (immunoblot) and immunohistochemistry (IHC). By comparing the staining from the blocked antibody versus the antibody alone, one can see which staining is specific; Specific binding will be absent from the western blot or immunostaining performed with the neutralized antibody.

Format and storage :

Synthetic peptide was lyophilized with 100% acetonitrile and is supplied as a powder. Reconstitute with 0.1 ml DI water for a final concentration of 10 mg/ml.The purity is >90%,tested by HPLC and MS.Storage Maintain refrigerated at 2-8°C for up to 6 months. For long term storage store at -20°C.

Precautions :

This product is for research use only. Not for use in diagnostic or therapeutic procedures.

Chicken
100%
Rabbit
100%
Pig
100%
Dog
100%
Sheep
100%
Horse
100%
Bovine
100%
Xenopus
90%
Zebrafish
0%
High similarity Medium similarity Low similarity No similarity
IMPORTANT: For western blots, incubate membrane with diluted antibody in 5% w/v milk , 1X TBS, 0.1% Tween®20 at 4°C with gentle shaking, overnight.

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