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  1. Link

    enos-antibody-m221-ab76198.pdf

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Neuroscience Neurotransmission Nitric Oxide NOS
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Anti-eNOS antibody [M221] (ab76198)

  • Datasheet
  • SDS
Reviews (14)Q&A (1)References (108)

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Western blot - Anti-eNOS antibody [M221] (ab76198)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eNOS antibody [M221] (ab76198)
  • Immunocytochemistry/ Immunofluorescence - Anti-eNOS antibody [M221] (ab76198)
  • Flow Cytometry (Intracellular) - Anti-eNOS antibody [M221] (ab76198)
  • Western blot - Anti-eNOS antibody [M221] (ab76198)

Key features and details

  • Mouse monoclonal [M221] to eNOS
  • Suitable for: IHC-P, ICC/IF, WB, Flow Cyt (Intra)
  • Reacts with: Mouse, Human
  • Isotype: IgG1

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Recombinant Human eNOS protein (ab112329)
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Overview

  • Product name

    Anti-eNOS antibody [M221]
    See all eNOS primary antibodies
  • Description

    Mouse monoclonal [M221] to eNOS
  • Host species

    Mouse
  • Specificity

    ab76198 is not predicted to react with other NOS family members due to low homology.

    This antibody detects eNOS in mouse and but at a lower intensity than in human. If you are working in mouse, we would recommend using no more than 1% milk as the blocking agent for optimal signal strength.

  • Tested applications

    Suitable for: IHC-P, ICC/IF, WB, Flow Cyt (Intra)more details
  • Species reactivity

    Reacts with: Mouse, Human
  • Immunogen

    Recombinant fragment corresponding to Mouse eNOS aa 1024-1202 (C terminal).
    Database link: P70313

  • Positive control

    • Human umbilical vein endothelial cells untreated and treated with lambda phosphatase. Mouse placenta lysate. Huvec lysate. IHC-P: FFPE human normal placenta tissue sections.
  • General notes

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.

    If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at -20°C. Stable for 12 months at -20°C.
  • Storage buffer

    Preservative: 0.05% Sodium azide
    Constituents: 0.1% BSA, 50% Glycerol, PBS
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    M221
  • Isotype

    IgG1
  • Research areas

    • Neuroscience
    • Neurotransmission
    • Nitric Oxide
    • NOS
    • Cardiovascular
    • Blood
    • Platelets
    • Cell Biology
    • Other Antibodies
    • Oxidative Stress
    • Cardiovascular
    • Vasculature
    • Vasodilation
    • Nitric oxide associated
    • Cancer
    • Cancer Metabolism
    • Response to hypoxia
    • Metabolism
    • Pathways and Processes
    • Metabolism processes
    • Hypoxia
    • Metabolism
    • Pathways and Processes
    • Redox metabolism
    • Oxidative stress
    • Metabolism
    • Types of disease
    • Cancer

Associated products

  • Compatible Secondaries

    • Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) (ab150113)
    • Goat Anti-Mouse IgG H&L (DyLight® 488) preadsorbed (ab96879)
  • Isotype control

    • Mouse IgG1, kappa monoclonal [15-6E10A7] - Isotype Control (ab170190)
    • Mouse IgG1, Kappa Monoclonal [B11/6] - Isotype Control (ab91353)
  • Recombinant Protein

    • Recombinant Human eNOS protein (ab112329)
  • Related Products

    • Prestained Protein Ladder – Broad molecular weight (10-245 kDa) (ab116028)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab76198 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
IHC-P
1/1000. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
ICC/IF (1)
Use at an assay dependent concentration.
WB (11)
1/500 - 1/1000. Predicted molecular weight: 133 kDa.
Flow Cyt (Intra)
Use 2µg for 106 cells.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

Notes
IHC-P
1/1000. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
ICC/IF
Use at an assay dependent concentration.
WB
1/500 - 1/1000. Predicted molecular weight: 133 kDa.
Flow Cyt (Intra)
Use 2µg for 106 cells.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

Target

  • Function

    Produces nitric oxide (NO) which is implicated in vascular smooth muscle relaxation through a cGMP-mediated signal transduction pathway. NO mediates vascular endothelial growth factor (VEGF)-induced angiogenesis in coronary vessels and promotes blood clotting through the activation of platelets.
    Isoform eNOS13C: Lacks eNOS activity, dominant-negative form that may down-regulate eNOS activity by forming heterodimers with isoform 1.
  • Tissue specificity

    Platelets, placenta, liver and kidney.
  • Involvement in disease

    Variation in NOS3 seem to be associated with susceptibility to coronary spasm.
  • Sequence similarities

    Belongs to the NOS family.
    Contains 1 FAD-binding FR-type domain.
    Contains 1 flavodoxin-like domain.
  • Post-translational
    modifications

    Phosphorylation by AMPK at Ser-1177 in the presence of Ca(2+)-calmodulin (CaM) activates activity. In absence of Ca(2+)-calmodulin, AMPK also phosphorylates Thr-495, resulting in inhibition of activity (By similarity). Phosphorylation of Ser-114 by CDK5 reduces activity.
  • Cellular localization

    Cell membrane. Membrane, caveola. Cytoplasm, cytoskeleton. Golgi apparatus. Specifically associates with actin cytoskeleton in the G2 phase of the cell cycle and which is favored by interaction with NOSIP and results in a reduced enzymatic activity.
  • Target information above from: UniProt accession P29474 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 4846 Human
    • Entrez Gene: 18127 Mouse
    • Omim: 163729 Human
    • SwissProt: P29474 Human
    • SwissProt: P70313 Mouse
    • Unigene: 647092 Human
    • Unigene: 258415 Mouse
    • Alternative names

      • cNOS antibody
      • Constitutive NOS antibody
      • EC NOS antibody
      • EC-NOS antibody
      • ecNOS antibody
      • Endothelial nitric oxidase synthase antibody
      • Endothelial nitric oxide synthase antibody
      • Endothelial nitric oxide synthase 3 antibody
      • Endothelial NOS antibody
      • eNOS antibody
      • Nitric oxide synthase 3 (endothelial cell) antibody
      • Nitric oxide synthase 3 antibody
      • Nitric oxide synthase 3 endothelial cell antibody
      • Nitric oxide synthase endothelial antibody
      • Nitric oxide synthase, endothelial antibody
      • NOS 3 antibody
      • NOS III antibody
      • NOS type III antibody
      • NOS3 antibody
      • NOS3_HUMAN antibody
      • NOSIII antibody
      see all

    Images

    • Western blot - Anti-eNOS antibody [M221] (ab76198)
      Western blot - Anti-eNOS antibody [M221] (ab76198)
      All lanes : Anti-eNOS antibody [M221] (ab76198) at 1/500 dilution

      Lane 1 : Huvec cell lysates
      Lane 2 : Mouse placenta lysates

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Goat anti Mouse IR680 at 1/10000 dilution

      Predicted band size: 133 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 using Licor blocking buffer before being incubated with ab76198 overnight at 4°C. Antibody binding was detected using Goat anti Mouse IR680 at a 1:10,000 dilution for 1hr at room temperature and then imaged using the Licor Odyssey CLx.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eNOS antibody [M221] (ab76198)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eNOS antibody [M221] (ab76198)

      IHC image of eNOS staining in human normal placenta*, performed on a Leica Bond™ 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 ab76198, 1/1000 dilution, 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.

      *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

    • Immunocytochemistry/ Immunofluorescence - Anti-eNOS antibody [M221] (ab76198)
      Immunocytochemistry/ Immunofluorescence - Anti-eNOS antibody [M221] (ab76198)

      ab76198 staining eNOS in human umbilical vein endothelial cells. Cells with fixed with paraformaldehyde.

       

    • Flow Cytometry (Intracellular) - Anti-eNOS antibody [M221] (ab76198)
      Flow Cytometry (Intracellular) - Anti-eNOS antibody [M221] (ab76198)
      Overlay histogram showing HEK293 cells stained with ab76198 (red line). The cells were fixed with methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab76198, 2µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a slightly decreased signal in HEK293 cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

    • Western blot - Anti-eNOS antibody [M221] (ab76198)
      Western blot - Anti-eNOS antibody [M221] (ab76198)
      All lanes : Anti-eNOS antibody [M221] (ab76198) at 1/1000 dilution

      Lane 1 : human umbilical vein endothelial
      cells, untreated
      Lane 2 : human umbilical vein endothelial
      cells, treated with lambda phosphatase

      Predicted band size: 133 kDa
      Observed band size: 140 kDa why is the actual band size different from the predicted?

    Protocols

    • Flow cytometry protocols
    • Immunohistochemistry protocols
    • Immunocytochemistry & immunofluorescence protocols
    • Western blot protocols

    Click here to view the general protocols

    Datasheets and documents

    • SDS download

    • Datasheet download

      Download

    References (108)

    Publishing research using ab76198? Please let us know so that we can cite the reference in this datasheet.

    ab76198 has been referenced in 108 publications.

    • Wee CL  et al. Calcitriol Supplementation Ameliorates Microvascular Endothelial Dysfunction in Vitamin D-Deficient Diabetic Rats by Upregulating the Vascular eNOS Protein Expression and Reducing Oxidative Stress. Oxid Med Cell Longev 2021:3109294 (2021). PubMed: 33623633
    • Liang L  et al. Exosomes derived from miR-301a-3p-overexpressing adipose-derived mesenchymal stem cells reverse hypoxia-induced erectile dysfunction in rat models. Stem Cell Res Ther 12:87 (2021). PubMed: 33494812
    • Wang JS  et al. Potential mechanism of Achyranthis bidentatae radix plus semen vaccariae granules in the treatment of diabetes mellitus-induced erectile dysfunction in rats utilizing combined experimental model and network pharmacology. Pharm Biol 59:547-556 (2021). PubMed: 33962551
    • Gan YR  et al. Dickkopf-1/cysteine-rich angiogenic inducer 61 axis mediates palmitic acid-induced inflammation and apoptosis of vascular endothelial cells. Mol Med Rep 23:N/A (2021). PubMed: 33300071
    • Lee-Huang S  et al. Endothelial Nitric Oxide Synthase Knockdown in Human Stem Cells Impacts Mitochondrial Biogenesis and Adipogenesis: Live-Cell Real-Time Fluorescence Imaging. J Clin Med 10:N/A (2021). PubMed: 33562279
    View all Publications for this product

    Customer reviews and Q&As

    Show All Reviews Q&A
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    1-10 of 15 Abreviews or Q&A

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-eNOS antibody [M221]

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Rabbit Tissue sections (Lung from neonatal rabbit)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citrate pH6
    Permeabilization
    No
    Specification
    Lung from neonatal rabbit
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 24°C
    Fixative
    Formaldehyde
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Rebeca Figueira

    Verified customer

    Submitted Mar 10 2021

    Western blot abreview for Anti-eNOS antibody [M221]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Rat Tissue lysate - other (Heart)
    Gel Running Conditions
    Reduced Denaturing (NuPAGE 4-12% Bis-Tris)
    Loading amount
    100 µg
    Specification
    Heart
    Blocking step
    Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Jan 06 2020

    Immunocytochemistry/ Immunofluorescence abreview for Anti-eNOS antibody [M221]

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (HUVEC)
    Permeabilization
    Yes - 0.1% Triton-X 100
    Specification
    HUVEC
    Blocking step
    BSA as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: 25°C
    Fixative
    10% Formalin
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Oct 16 2019

    Western blot abreview for Anti-eNOS antibody [M221]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Mouse Tissue lysate - whole (Kidney)
    Gel Running Conditions
    Reduced Denaturing (4-12%)
    Loading amount
    40 µg
    Specification
    Kidney
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    DR. Wesley Konsavage

    Verified customer

    Submitted Dec 21 2016

    Western blot abreview for Anti-eNOS antibody [M221]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Rat Tissue lysate - whole (Skeletal muscle)
    Gel Running Conditions
    Reduced Denaturing (8%)
    Loading amount
    20 µg
    Specification
    Skeletal muscle
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Feb 09 2016

    Western blot abreview for Anti-eNOS antibody [M221]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Rat Tissue lysate - whole (Heart)
    Gel Running Conditions
    Reduced Denaturing (10%)
    Loading amount
    50 µg
    Specification
    Heart
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Jan 26 2016

    Western blot abreview for Anti-eNOS antibody [M221]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Rat Tissue lysate - whole (skeletal muscle (plantaris))
    Gel Running Conditions
    Reduced Denaturing (8%)
    Loading amount
    40 µg
    Specification
    skeletal muscle (plantaris)
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Dec 04 2015

    Western blot abreview for Anti-eNOS antibody [M221]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Loading amount
    20 µg
    Gel Running Conditions
    Reduced Denaturing (8%)
    Sample
    Rat Tissue lysate - whole (Skeletal muscle)
    Specification
    Skeletal muscle
    Blocking step
    Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Jan 28 2015

    Western blot abreview for Anti-eNOS antibody [M221]

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Loading amount
    20 µg
    Gel Running Conditions
    Reduced Denaturing
    Sample
    Rat Tissue lysate - whole (Rat brain extract)
    Specification
    Rat brain extract
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Jun 23 2014

    Western blot abreview for Anti-eNOS antibody [M221]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Loading amount
    50 µg
    Gel Running Conditions
    Reduced Denaturing (8% Gel)
    Sample
    Human Tissue lysate - whole (Vastus lateralis muscle)
    Specification
    Vastus lateralis muscle
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Feb 18 2014

    1-10 of 15 Abreviews or Q&A

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