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    vcam1-antibody-epr5047-low-endotoxin-azide-free-ab215380.pdf

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Signal Transduction Cytoskeleton / ECM Cell Adhesion Cell Adhesion Molecules Vascular
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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

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  • Certificate of Compliance
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Western blot - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
  • Flow Cytometry (Intracellular) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
  • Immunocytochemistry/ Immunofluorescence - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
  • Immunoprecipitation - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
  • Immunocytochemistry/ Immunofluorescence - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
  • Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR5047] to VCAM1 - Low endotoxin, Azide free
  • Suitable for: Flow Cyt (Intra), WB, IP, IHC-P, ICC/IF
  • Knockout validated
  • Reacts with: Mouse, Human

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Overview

  • Product name

    Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free
    See all VCAM1 primary antibodies
  • Description

    Rabbit monoclonal [EPR5047] to VCAM1 - Low endotoxin, Azide free
  • Host species

    Rabbit
  • Tested applications

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

    Reacts with: Mouse, Human
    Predicted to work with: Rat
  • Immunogen

    Full length protein. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: Human fetal liver, HuT 78, NIH 3T3, Mouse brain, Mouse kidney, Mouse spleen, Rat brain, Rat kidney and Rat spleen lysates; Wild-type HUVEC and A549 TNF-a treated (10 ng/mL, 16h) cell lysates. IHC-P: Human and Mouse spleen FFPE tissue sections. Flow Cyt (intra): K562 cells. ICC/IF: HUVEC TNF-a treated (10 ng/mL, 16h); K562 cells.
  • General notes

    ab215380 is the carrier-free version of ab134047.

    Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.

    This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

    Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

    Our Low endotoxin, azide-free formats have low endotoxin level (≤ 1 EU/ml, determined by the LAL assay) and are free from azide, to achieve consistent experimental results in functional assays.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C. Do Not Freeze.
  • Storage buffer

    pH: 7.2
    Constituent: PBS
  • Carrier free

    Yes
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR5047
  • Isotype

    IgG
  • Research areas

    • Signal Transduction
    • Cytoskeleton / ECM
    • Cell Adhesion
    • Cell Adhesion Molecules
    • Vascular
    • Signal Transduction
    • Cytoskeleton / ECM
    • Cell Adhesion
    • Cell Adhesion Molecules
    • Endothelial
    • Neuroscience
    • Neurology process
    • Neural Signal Transduction
    • Stem Cells
    • Mesenchymal Stem Cells
    • Surface Molecules
    • Microbiology
    • Organism
    • Virus
    • RNA Virus
    • ssRNA positive strand virus
    • SARS Coronavirus
    • Cancer
    • Invasion/microenvironment
    • ECM
    • Cell adhesion
    • Other
    • Cardiovascular
    • Atherosclerosis
    • Vascular Inflammation
    • Leukocyte recruitment
    • Cell adhesion molecules
    • Kits/ Lysates/ Other
    • Kits
    • ELISA Kits
    • ELISA Kits
    • Adhesion molecules ELISA kits
    • Kits/ Lysates/ Other
    • Kits
    • ELISA Kits
    • ELISA Kits
    • Atherosclerotic proteins ELISA kits
    • Cardiovascular
    • Angiogenesis
    • Endothelial Cell Markers

Associated products

  • Alternative Versions

    • Anti-VCAM1 antibody [EPR5047] (ab134047)
    • Alexa Fluor® 647 Anti-VCAM1 antibody [EPR5047] (ab194319)
    • HRP Anti-VCAM1 antibody [EPR5047] (ab195540)
    • PE Anti-VCAM1 antibody [EPR5047] (ab223981)
    • FITC Anti-VCAM1 antibody [EPR5047] (ab223982)
    • APC Anti-VCAM1 antibody [EPR5047] (ab223983)
  • Conjugation kits

    • PE / R-Phycoerythrin Conjugation Kit - Lightning-Link® (ab102918)
    • APC Conjugation Kit - Lightning-Link® (ab201807)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Positive Controls

    • Rat brain normal tissue lysate - membrane extract (ab29473)
  • Related Products

    • Human VCAM1 knockout A549 cell line (ab273758)
    • Human VCAM1 knockout A549 cell lysate (ab275504)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab215380 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
Flow Cyt (Intra)
Use at an assay dependent concentration.
WB
Use at an assay dependent concentration.
IP
Use at an assay dependent concentration.
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocol.

ICC/IF
Use at an assay dependent concentration.
Notes
Flow Cyt (Intra)
Use at an assay dependent concentration.
WB
Use at an assay dependent concentration.
IP
Use at an assay dependent concentration.
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocol.

ICC/IF
Use at an assay dependent concentration.

Target

  • Function

    Important in cell-cell recognition. Appears to function in leukocyte-endothelial cell adhesion. Interacts with the beta-1 integrin VLA4 on leukocytes, and mediates both adhesion and signal transduction. The VCAM1/VLA4 interaction may play a pathophysiologic role both in immune responses and in leukocyte emigration to sites of inflammation.
  • Tissue specificity

    Expressed on inflammed vascular endothelium, as well as on macrophage-like and dendritic cell types in both normal and inflammed tissue.
  • Sequence similarities

    Contains 7 Ig-like C2-type (immunoglobulin-like) domains.
  • Domain

    Either the first or the fourth Ig-like C2-type domain is required for VLA4-dependent cell adhesion.
  • Post-translational
    modifications

    Sialoglycoprotein.
  • Cellular localization

    Membrane.
  • Target information above from: UniProt accession P19320 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 7412 Human
    • Entrez Gene: 22329 Mouse
    • Entrez Gene: 25361 Rat
    • GenBank: NP_001069.1 Human
    • GenBank: NP_001186763.1 Human
    • Omim: 192225 Human
    • SwissProt: P19320 Human
    • SwissProt: P29533 Mouse
    • SwissProt: P29534 Rat
    • Unigene: 109225 Human
    • Unigene: 440909 Mouse
    • Unigene: 76649 Mouse
    • Unigene: 11267 Rat
    see all
  • Alternative names

    • CD106 antibody
    • CD106 Antigen antibody
    • INCAM 100 antibody
    • INCAM-100 antibody
    • L1CAM antibody
    • MGC99561 antibody
    • V-CAM 1 antibody
    • Vascular Cell Adhesion Molecule 1 antibody
    • Vascular cell adhesion protein 1 antibody
    • VCAM 1 antibody
    • VCAM-1 antibody
    • VCAM1 antibody
    • VCAM1_HUMAN antibody
    see all

Images

  • Western blot - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    Western blot - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    All lanes : Anti-VCAM1 antibody [EPR5047] (ab134047) at 1/2000 dilution

    Lane 1 : Wild-type A549 cell lysate
    Lane 2 : Wild-type A549 TNF-a treated (10 ng/mL, 16h) cell lysate
    Lane 3 : VCAM1 knockout A549 cell lysate
    Lane 4 : VCAM1 knockout A549 TNF-a treated (10 ng/mL, 16h) cell lysate
    Lane 5 : HUVEC cell lysate
    Lane 6 : HUVEC TNF-a treated (16 ng/mL, 16h) cell lysate

    Lysates/proteins at 30 µg per lane.

    Performed under reducing conditions.

    Observed band size: 105 kDa why is the actual band size different from the predicted?



    This data was developed using the same antibody clone in a different buffer formulation (ab134047).

    Lanes 1 - 6: Merged signal (red and green). Green - ab134047 observed at 105 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

    ab134047 was shown to react with VCAM1 in wild-type A549 cells in Western blot with loss of signal observed in VCAM1 knockout sample. Wild-type A549 and VCAM1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab134047 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with HRP conjugated Goat anti-Rabbit (H+L) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

    IHC image of VCAM1 staining in Mouse spleen formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol B. 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 unpurified ab134047, 1/200 dilution, for 15 mins at room temperature. A goat anti-rabbit biotinylated secondary antibody was used to detect the primary, and visualized using an HRP conjugated ABC 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.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134047).

  • Flow Cytometry (Intracellular) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    Flow Cytometry (Intracellular) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

    Intracellular Flow Cytometry analysis of K562 (human chronic myelogenous leukemia) cells labeling VCAM1 with purified ab134047 at 1/40 dilution (10ug/mL) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor®488) at 1/2000 dilution was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) was used as the unlabeled control. 

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134047). 

     

     

  • Immunocytochemistry/ Immunofluorescence - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    Immunocytochemistry/ Immunofluorescence - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

    ab134047 staining VCAM1 in HUVEC cells treated with TNF-α (ab55237) at 10 ng/ml for 16 hours. The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab134047 at 2μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 647) (ab150119) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
    Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).
    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134047).

  • Immunoprecipitation - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    Immunoprecipitation - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

    VCAM1 was immunoprecipitated from Human fetal liver lysate with ab134047 at 1/110 dilution.

    Western blot was performed from the immunoprecipitate using ab134047 at 1/1000 dilution.

    VeriBlot for IP secondary antibody (Peroxidase conjugated),was used as secondary antibody at 1/1000 dilution.

    Lane 1:Human fetal liver lysate

    Blocking and dilution buffer:5% NFDM/TBST

    Exposure time: 1 second

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134047).

  • Immunocytochemistry/ Immunofluorescence - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    Immunocytochemistry/ Immunofluorescence - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

    Immunofluorescent staining of K562 cells (fixed in 4% PFA, permeabilized with 0.1% Triton X 100) using purified ab134047 at a dilution of 1/250. An Alexa Fluor® 488 goat anti-rabbit antibody was used as the secondary at a dilution of 1/500 and the cells were counter stained with DAPI. The negative control is shown in the bottom right hand panel - for the negative control, Alex Fluor® 594 goat anti-mouse was used at a dilution of 1/500.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134047).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

    This IHC data was generated using the same anti-VCAM1 antibody clone, EPR5047, in a different buffer formulation (cat# ab134047).

    IHC image of VCAM1 staining in Human spleen formalin fixed paraffin embedded tissue section, 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 unpurified ab134047, 1/200 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.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

    This IHC data was generated using the same anti-VCAM1 antibody clone, EPR5047, in a different buffer formulation (cat# ab134047).

    Immunohistochemical staining of paraffin embedded human tonsil with purified ab134047 at a dilution of 1/500. A pre-diluted HRP polymer for rabbit/mouse IgG was used as the secondary antibody and the sample was counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

  • Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)
    Anti-VCAM1 antibody [EPR5047] - Low endotoxin, Azide free (ab215380)

Protocols

  • Immunoprecipitation protocols
  • Immunohistochemistry protocols
  • Immunocytochemistry & immunofluorescence protocols

Click here to view the general protocols

Datasheets and documents

  • Datasheet download

    Download

Certificate of Compliance

To download a Certificate of Compliance, please enter your Lot number below:

References (9)

Publishing research using ab215380? Please let us know so that we can cite the reference in this datasheet.

ab215380 has been referenced in 9 publications.

  • Sun C  et al. ADAM17-regulated CX3CL1 expression produced by bone marrow endothelial cells promotes spinal metastasis from hepatocellular carcinoma. Int J Oncol 57:249-263 (2020). PubMed: 32319605
  • Qureshi AW  et al. Ageing enhances the shedding of splenocyte microvesicles with endothelial pro-senescent effect that is prevented by a short-term intake of omega-3 PUFA EPA:DHA 6:1. Biochem Pharmacol 173:113734 (2020). PubMed: 31811867
  • Pang X & Hou X Synergistic protective effect of FTY720 and vitamin E against simulated cerebral ischemia in vitro. Mol Med Rep : (2017). PubMed: 28498446
  • Wu F  et al. CXCR2 is essential for cerebral endothelial activation and leukocyte recruitment during neuroinflammation. J Neuroinflammation 12:98 (2015). WB ; Mouse . PubMed: 25990934
  • Coon BG  et al. Intramembrane binding of VE-cadherin to VEGFR2 and VEGFR3 assembles the endothelial mechanosensory complex. J Cell Biol 208:975-86 (2015). PubMed: 25800053
  • Wu M  et al. microRNA-23b regulates the expression of inflammatory factors in vascular endothelial cells during sepsis. Exp Ther Med 9:1125-1132 (2015). WB ; Human . PubMed: 25780398
  • Nakayama J  et al. Decreased peritoneal ovarian cancer growth in mice lacking expression of lipid phosphate phosphohydrolase 1. PLoS One 10:e0120071 (2015). IHC . PubMed: 25769037
  • Gregory EK  et al. Periadventitial atRA citrate-based polyester membranes reduce neointimal hyperplasia and restenosis after carotid injury in rats. Am J Physiol Heart Circ Physiol 307:H1419-29 (2014). PubMed: 25239800
  • Florea V  et al. c-Myc Is Essential to Prevent Endothelial Pro-Inflammatory Senescent Phenotype. PLoS One 8:e73146 (2013). WB . PubMed: 24039874

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