Anti-Pyruvate Dehydrogenase E1-alpha subunit (phospho S293) 抗体 (ab92696)

製品の概要

  • 製品名
    Anti-Pyruvate Dehydrogenase E1-alpha subunit (phospho S293) antibody
    Pyruvate Dehydrogenase E1-alpha subunit 一次抗体 製品一覧
  • 製品の詳細
    Rabbit polyclonal to Pyruvate Dehydrogenase E1-alpha subunit (phospho S293)
  • アプリケーション
    適用あり: WB, ICC/IF, IHC-Pmore details
  • 種交差性
    交差種: Mouse, Human
    交差が予測される動物種: Rat, Cow, Pig, Orangutan
  • 免疫原

    Synthetic peptide conjugated to KLH derived from within residues 250 - 350 of Human Pyruvate Dehydrogenase E1-alpha subunit, phosphorylated at S293.

  • ポジティブ・コントロール
    • This antibody gave a positive signal in Human liver tissue lysate as well as the following whole cell lysates: HeLa; HEK293; HepG2. This antibody gave a positive result in IHC in the following FFPE tissue: Human lung adenocarcinoma. ICC-IF: HepG2 cells.

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab92696 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

アプリケーション Abreviews 特記事項
WB Use a concentration of 1 µg/ml. Detects a band of approximately 43 kDa (predicted molecular weight: 43 kDa).
ICC/IF Use a concentration of 1 - 5 µg/ml.
IHC-P Use a concentration of 1 µg/ml.

ターゲット情報

  • 機能
    The pyruvate dehydrogenase complex catalyzes the overall conversion of pyruvate to acetyl-CoA and CO(2). It contains multiple copies of three enzymatic components: pyruvate dehydrogenase (E1), dihydrolipoamide acetyltransferase (E2) and lipoamide dehydrogenase (E3).
  • 組織特異性
    Ubiquitous.
  • 関連疾患
    Defects in PDHA1 are a cause of pyruvate decarboxylase E1 component deficiency (PDHE1 deficiency) [MIM:312170]. PDHE1 deficiency is the most common enzyme defect in patients with primary lactic acidosis. It is associated with variable clinical phenotypes ranging from neonatal death to prolonged survival complicated by developmental delay, seizures, ataxia, apnea, and in some cases to an X-linked form of Leigh syndrome (X-LS).
    Defects in PDHA1 are the cause of X-linked Leigh syndrome (X-LS) [MIM:308930]. X-LS is an early-onset progressive neurodegenerative disorder with a characteristic neuropathology consisting of focal, bilateral lesions in one or more areas of the central nervous system, including the brainstem, thalamus, basal ganglia, cerebellum, and spinal cord. The lesions are areas of demyelination, gliosis, necrosis, spongiosis, or capillary proliferation. Clinical symptoms depend on which areas of the central nervous system are involved. The most common underlying cause is a defect in oxidative phosphorylation. LS may be a feature of a deficiency of any of the mitochondrial respiratory chain complexes.
  • 細胞内局在
    Mitochondrion matrix.
  • Information by UniProt
  • 参照データベース
  • 別名
    • ODPA_HUMAN antibody
    • PDH antibody
    • PDHA antibody
    • PDHA1 antibody
    • PDHCE1A antibody
    • PDHE1 A type I antibody
    • PDHE1-A type I antibody
    • PHE1A antibody
    • Pyruvate Dehydrogenase (lipoamide) alpha 1 antibody
    • Pyruvate dehydrogenase complex, E1 alpha polypeptide 1 antibody
    • Pyruvate Dehydrogenase E1 alpha antibody
    • Pyruvate dehydrogenase E1 component subunit alpha, somatic form, mitochondrial antibody
    see all

画像

  • All lanes : Anti-Pyruvate Dehydrogenase E1-alpha subunit (phospho S293) antibody (ab92696) at 1 µg/ml

    Lane 1 : HeLa Whole Cell Lysate + Calyculin A (30 nM for 20 min)
    Lane 2 : HeLa Whole Cell Lysate + Calyculin A (30 nM for 20 min)

    Lysates/proteins at 20 µg per lane.

    Secondary
    goat anti-rabbit (green) and goat anti-mouse (red) at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size : 43 kDa

    This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour and then treated with either buffer (lane 1) or alkaline phosphatase (lane 2), before being incubated with ab92696 overnight at 4°C. Antibody binding was detected using IR-labelled goat anti-rabbit (green) and goat anti-mouse (Red) at 1:10,000 dilution for one hour at room temperature before imaging.

     

  • ab92696 stained in HepG2 cells. Cells were fixed with 100% paraformaldehyde (5 min) at room temperature and incubated with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% triton for 1h at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab92696 at 5 µg/ml and ab7291 (Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control) at 1/1000 dilution overnight at +4°C. The secondary antibodies were ab150120 (pseudo-colored red) and ab150081 (colored green) used at 1 ug/ml for 1hour at room temperature. DAPI was used to stain the cell nuclei (colored blue) at a concentration of 1.43 µM for 1hour at room temperature

  • IHC image of Pyruvate Dehydrogenase E1-alpha subunit (phospho S293) staining in Human lung adenocarcinoma formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab92696, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • Anti-Pyruvate Dehydrogenase E1-alpha subunit (phospho S293) antibody (ab92696) at 1/1000 dilution + Human vascular smooth muscle cell whole cell lysate at 25 µg

    Secondary
    Polyclonal goat anti-rabbit IRDye® 800CW at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size : 43 kDa
    Observed band size : 43 kDa


    Exposure time : 5 minutes

    This image is courtesy of an anonymous abreview.

    See Abreview

  • Serially diluted ab92696 was bound to immobilised Phospho peptide (S293) - or Control peptide (1 microgram x mL-1). The antibody was detected by HRP-labelled goat anti-rabbit IgG

  • ab92696 staining Pyruvate Dehydrogenase E1-alpha subunit (phospho S293) in Human HUVEC cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.1% Triton X-100 pH 7.4 for 5 minutes and blocked with 5% BSA for 20 minutes at room temperature. Samples were incubated with primary antibody (1/500 in PBS) for 1 hour . A CF488-conjugated Donkey anti-rabbit IgG polyclonal (1/500) was used as the secondary antibody.

    See Abreview

  • All lanes : Anti-Pyruvate Dehydrogenase E1-alpha subunit (phospho S293) antibody (ab92696) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate
    Lane 3 : HEK293 (Human embryonic kidney cell line) Whole Cell Lysate
    Lane 4 : Human liver tissue lysate - total protein (ab29889)
    Lane 5 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate with Modified blocking peptide at 1 µg/ml
    Lane 6 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate with Modified blocking peptide at 1 µg/ml
    Lane 7 : HEK293 (Human embryonic kidney cell line) Whole Cell Lysate with Modified blocking peptide at 1 µg/ml
    Lane 8 : Human liver tissue lysate - total protein (ab29889) with Modified blocking peptide at 1 µg/ml
    Lane 9 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate with Non-modified blocking peptide at 1 µg/ml
    Lane 10 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate with Non-modified blocking peptide at 1 µg/ml
    Lane 11 : HEK293 (Human embryonic kidney cell line) Whole Cell Lysate with Non-modified blocking peptide at 1 µg/ml
    Lane 12 : Human liver tissue lysate - total protein (ab29889) with Non-modified blocking peptide at 1 µg/ml

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 43 kDa
    Observed band size : 43 kDa
    Additional bands at : 34 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 2 minutes

参考文献

This product has been referenced in:
  • Ward NP  et al. Complex I inhibition augments dichloroacetate cytotoxicity through enhancing oxidative stress in VM-M3 glioblastoma cells. PLoS One 12:e0180061 (2017). WB . Read more (PubMed: 28644886) »
  • Ofori JK  et al. Elevated miR-130a/miR130b/miR-152 expression reduces intracellular ATP levels in the pancreatic beta cell. Sci Rep 7:44986 (2017). Read more (PubMed: 28332581) »

See all 13 Publications for this product

レビューと Q&A

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

Abcam user community

Verified customer

投稿 Jul 25 2017

Application
Western blot
Sample
Zebrafish Tissue lysate - whole (Brain, retina)
Gel Running Conditions
Reduced Denaturing (10% gel)
Loading amount
20 µg
Specification
Brain, retina
Blocking step
Licor Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 4°C
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Abcam user community

Verified customer

投稿 Jun 20 2017

Application
Western blot
Sample
Mouse Tissue lysate - whole (Brain, liver, retina, RPE)
Gel Running Conditions
Reduced Denaturing (10% gel)
Loading amount
20 µg
Specification
Brain, liver, retina, RPE
Blocking step
Licor Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C
Username

Abcam user community

Verified customer

投稿 Jun 20 2017

Application
Western blot
Sample
Human Tissue lysate - whole (HEK cells, ARPE19 cells)
Gel Running Conditions
Reduced Denaturing (10% gel)
Loading amount
20 µg
Specification
HEK cells, ARPE19 cells
Blocking step
Licor Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C
Username

Abcam user community

Verified customer

投稿 Jun 20 2017

Application
Western blot
Sample
Human Cell lysate - whole cell (Vascular Smooth Muscle Cell)
Gel Running Conditions
Reduced Denaturing
Loading amount
25 µg
Specification
Vascular Smooth Muscle Cell
Blocking step
Milk as blocking agent for 3 hour(s) and 0 minute(s) · Concentration: 5µg/mL · Temperature: 21°C
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投稿 Jul 20 2016

Application
Western blot
Sample
Sheep Tissue lysate - whole (liver)
Gel Running Conditions
Reduced Denaturing
Loading amount
30 µg
Specification
liver
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Jennifer Bruce

Verified customer

投稿 Jun 15 2015

Application
Western blot
Loading amount
20 µg
Gel Running Conditions
Reduced Denaturing (10%)
Sample
Human Cell lysate - whole cell (Human lung adenocarcinoma epithelial cell line)
Specification
Human lung adenocarcinoma epithelial cell line
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Dr. Stamatia Pouliliou

Verified customer

投稿 Mar 23 2015

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
BSA as blocking agent for 20 minute(s) · Concentration: 5%
Sample
Human Cell (HuVEC)
Specification
HuVEC
Permeabilization
Yes - 0.1% v/v Triton X-100 pH 7.4 for 5 min at RT
Fixative
Paraformaldehyde
Username

Dr. Dimitra Kalamida

Verified customer

投稿 Nov 18 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Tissue lysate - whole (Heart, left ventricle)
Loading amount
40 µg
Specification
Heart, left ventricle
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Dawn Bowles

Verified customer

投稿 Dec 27 2012

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - other (HeLa and U2OS: Purified mitochondria lysate from 9)
Loading amount
3e+006 cells
Specification
HeLa and U2OS: Purified mitochondria lysate from 9
Gel Running Conditions
Reduced Denaturing (8% polyacrylamide gel)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

投稿 Aug 09 2012

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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