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    vimentin-antibody-v9-bsa-and-azide-free-ab223871.pdf

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Validated using a knockout cell line

Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

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Western blot - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
  • Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
  • Flow Cytometry - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
  • Western blot - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
  • Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
  • Flow Cytometry - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
  • Flow Cytometry - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

Key features and details

  • Mouse monoclonal [V9] to Vimentin - BSA and Azide free
  • Suitable for: IHC-Fr, WB, IHC-P, ICC/IF, Flow Cyt, IHC-FoFr
  • Knockout validated
  • Reacts with: Rat, Human
  • Isotype: IgG1

Conjugates logo Related conjugates and formulations

Alexa Fluor® 488 Alexa Fluor® 647 HRP

You may also be interested in

Secondary
Product image
Goat Anti-Mouse IgG H&L (HRP) (ab205719)
Knockout
Product image
Human VIM (Vimentin) knockout HeLa cell line (ab255446)

View more associated products

Overview

  • Product name

    Anti-Vimentin antibody [V9] - BSA and Azide free
    See all Vimentin primary antibodies
  • Description

    Mouse monoclonal [V9] to Vimentin - BSA and Azide free
  • Host species

    Mouse
  • Tested applications

    Suitable for: IHC-Fr, WB, IHC-P, ICC/IF, Flow Cyt, IHC-FoFrmore details
  • Species reactivity

    Reacts with: Rat, Human
    Predicted to work with: Horse, Chicken, Cow, Cat, Dog, Pig
  • Immunogen

    Full length native protein (purified). This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: HeLa, U-2 OS, human tonsil, HAP1 and MOLT4 whole cell lysates. ICC/IF: HeLa cells. IHC-P: Human kidney FFPE tissue sections. Flow Cyt: MDA-MB-231, HAP1 and SV40LT-SMC cells.
  • General notes

    ab223871 is the carrier-free version of ab8069.

    This monoclonal antibody to vimentin has been knockout validated in ICC/IF. The expected staining was observed in wild type cells and no staining was seen in vimentin knockout cells.

    This antibody clone is manufactured by Abcam. If you require a custom buffer formulation or conjugation for your experiments, please contact orders@abcam.com.

    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.

    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 +4°C. Do Not Freeze.
  • Storage buffer

    Constituent: PBS
  • Carrier free

    Yes
  • Concentration information loading...
  • Purity

    IgG fraction
  • Clonality

    Monoclonal
  • Clone number

    V9
  • Isotype

    IgG1
  • Research areas

    • Neuroscience
    • Cell Type Marker
    • Neural Stem Cell marker
    • Signal Transduction
    • Cytoskeleton / ECM
    • Cytoskeleton
    • Intermediate Filaments
    • Class III
    • Vimentin
    • Neuroscience
    • Neurology process
    • Neurogenesis
    • Stem Cells
    • Lineage Markers
    • Endoderm
    • Stem Cells
    • Neural Stem Cells
    • Intracellular
    • Cancer
    • Tumor biomarkers
    • Other
    • Neuroscience
    • Neurology process
    • Neuroregeneration
    • Neuroregeneration
    • Developmental Biology
    • Lineage specification
    • Ectoderm
    • Neuroscience
    • Development
    • Neuroscience
    • Processes

Associated products

  • Alternative Versions

    • Alexa Fluor® 488 Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab195877)
    • Alexa Fluor® 647 Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab195878)
    • HRP Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab196602)
    • Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)
  • Compatible Secondaries

    • Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) (ab150113)
    • Goat Anti-Mouse IgG H&L (HRP) (ab205719)
    • Goat Anti-Mouse IgG H&L (DyLight® 488) preadsorbed (ab96879)
  • Isotype control

    • Mouse IgG1, kappa monoclonal [15-6E10A7] - Isotype Control (ab170190)
  • KO cell lines

    • Human VIM (Vimentin) knockout HeLa cell line (ab255446)
  • KO cell lysates

    • Human VIM (Vimentin) knockout HeLa cell lysate (ab263775)
  • Related Products

    • Human Vimentin Profiling ELISA Kit (ab173190)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab223871 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-Fr
Use at an assay dependent concentration.
WB
Use at an assay dependent concentration. Detects a band of approximately 57 kDa (predicted molecular weight: 54 kDa).
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
ICC/IF
Use at an assay dependent concentration.
Flow Cyt
Use at an assay dependent concentration.

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

IHC-FoFr
Use at an assay dependent concentration.
Notes
IHC-Fr
Use at an assay dependent concentration.
WB
Use at an assay dependent concentration. Detects a band of approximately 57 kDa (predicted molecular weight: 54 kDa).
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
ICC/IF
Use at an assay dependent concentration.
Flow Cyt
Use at an assay dependent concentration.

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

IHC-FoFr
Use at an assay dependent concentration.

Target

  • Function

    Vimentins are class-III intermediate filaments found in various non-epithelial cells, especially mesenchymal cells. Vimentin is attached to the nucleus, endoplasmic reticulum, and mitochondria, either laterally or terminally.
    Involved with LARP6 in the stabilization of type I collagen mRNAs for CO1A1 and CO1A2.
  • Tissue specificity

    Highly expressed in fibroblasts, some expression in T- and B-lymphocytes, and little or no expression in Burkitt's lymphoma cell lines. Expressed in many hormone-independent mammary carcinoma cell lines.
  • Involvement in disease

    Cataract 30
  • Sequence similarities

    Belongs to the intermediate filament family.
  • Domain

    The central alpha-helical coiled-coil rod region mediates elementary homodimerization.
    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.
  • Post-translational
    modifications

    Filament disassembly during mitosis is promoted by phosphorylation at Ser-55 as well as by nestin (By similarity). One of the most prominent phosphoproteins in various cells of mesenchymal origin. Phosphorylation is enhanced during cell division, at which time vimentin filaments are significantly reorganized. Phosphorylation by PKN1 inhibits the formation of filaments. Phosphorylated at Ser-56 by CDK5 during neutrophil secretion in the cytoplasm. Phosphorylated by STK33.
    O-glycosylated during cytokinesis at sites identical or close to phosphorylation sites, this interferes with the phosphorylation status.
    S-nitrosylation is induced by interferon-gamma and oxidatively-modified low-densitity lipoprotein (LDL(ox)) possibly implicating the iNOS-S100A8/9 transnitrosylase complex.
  • Cellular localization

    Cytoplasm.
  • Target information above from: UniProt accession P08670 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 420519 Chicken
    • Entrez Gene: 280955 Cow
    • Entrez Gene: 477991 Dog
    • Entrez Gene: 7431 Human
    • Entrez Gene: 100522394 Pig
    • Entrez Gene: 81818 Rat
    • Omim: 193060 Human
    • SwissProt: P09654 Chicken
    • SwissProt: P48616 Cow
    • SwissProt: P08670 Human
    • SwissProt: P02543 Pig
    • SwissProt: P31000 Rat
    • Unigene: 455493 Human
    • Unigene: 691131 Human
    • Unigene: 2710 Rat
    see all
  • Form

    Vimentin is found in connective tissue and in the cytoskeleton.
  • Alternative names

    • CTRCT30 antibody
    • Epididymis luminal protein 113 antibody
    • FLJ36605 antibody
    • HEL113 antibody
    • VIM antibody
    • VIME_HUMAN antibody
    • Vimentin antibody
    see all

Images

  • Western blot - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Western blot - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    All lanes : Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069) at 1 µg

    Lane 1 : U-2 OS cell lysate
    Lane 2 : Human tonsil cell lysate
    Lane 3 : Wild-type HeLa cell lysate
    Lane 4 : VIM knockout HeLa cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 54 kDa
    Observed band size: 53 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 (ab8069).

    Lanes 1 - 4: Merged signal (red and green). Green - ab8069 observed at 53 kDa. Red - loading control, ab181602 observed at 37 kDa.

     ab8069 was shown to react with Vimentin in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab255446 (knockout cell lysate ab263775) was used. Wild-type and Vimentin knockout samples were subjected to SDS-PAGE. ab8069 and Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) were incubated overnight at 4°C at 1 µg/ml and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

     

     

  • Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

    This image was produced using the same antibody clone, but in a different formulation, ab8069.

    ab8069 staining Vimentin (colored green) in wild-type HAP1 cells (top panel) and VIM knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), 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 ab8069 at 0.5μg/ml and ab202272 (Rabbit monoclonal [EP1332Y] to alpha Tubulin (Alexa Fluor® 594) at 1/250 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with ab150117 (Goat secondary antibody to Mouse IgG (Alexa Fluor® 488) at 2 μg/ml (colored green). Nuclear DNA was labeled in blue with DAPI.

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

    This image was produced using the same antibody clone, but in a different formulation, ab8069.

    IHC image of ab8069 staining Vimentin in normal human kidney formalin-fixed paraffin-embedded tissue sections*, performed on a Leica Bond. 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 ab8069, 1/500 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 hematoxylin and mounted with DPX.

    No primary antibody was used in the negative control (shown on the inset).

    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

  • Flow Cytometry - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Flow Cytometry - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

    Flow cytometry overlay histogram showing wild-type HAP1 (green line) and VIM knockout HAP1 cells stained with ab223871 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab223871) (1x106 in 100 µl at 0.04 µg/ml) for 30 min at 22°C.

    The secondary antibody Goat anti-mouse IgG H&L (Alexa Fluor® 488, pre-adsorbed) (ab150117) was used at 1/2000 for 30 min at 22°C.

    Isotype control antibody was mouse IgG1&kappa (ab170190) used at the same concentration and conditions as the primary antibody (wild-type HAP1 - black line VIM knockout HAP1 - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).

    Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

    This antibody can also be used in in HAP1 cells fixed with 4% formaldehyde (10 min) / permeabilized with 0.1% PBS-Triton X-100 for 15 min used under the same conditions.

     

  • Western blot - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Western blot - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    All lanes : Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069) at 1 µg/ml

    Lane 1 : Wild-type HAP1 whole cell lysate
    Lane 2 : VIM (Vimentin) knockout HAP1 whole cell lysate
    Lane 3 : HeLa whole cell lysate
    Lane 4 : MOLT4 whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 54 kDa



    This image was produced using the same antibody clone, but in a different formulation, ab8069.

    Lanes 1 - 4: Merged signal (red and green). Green - ab8069 observed at 57 kDa. Red - loading control, ab181602, observed at 37 kDa.

    ab8069 was shown to specifically react with Vimentin in wild-type HAP1 cells as signal was lost in VIM (Vimentin) knockout cells. Wild-type and VIM (Vimentin) knockout samples were subjected to SDS-PAGE. Ab8069 and ab181602 (Rabbit anti-GAPDH loading control) were incubated overnight at 4°C at 1 μg/ml and 1/10000 dilution respectively. Blots were developed with goat anti-mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and goat anti-rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

  • Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

    This image was produced using the same antibody clone, but in a different formulation, ab8069.

    ab8069 staining Vimentin in HeLa cells. The cells were fixed with 4% formaldehyde (10min), 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 overnight at +4°C with ab8069 at 1/100 dilution (shown in green) and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 594), at 2µg/ml (shown in red). Nuclear DNA was labeled with DAPI (shown in blue).
    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

    This image was produced using the same antibody clone, but in a different formulation, ab8069.

    IHC image of Vimentin staining in human Hodgkin's lymphoma 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 ab8069, 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 hematoxylin and mounted with DPX.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

    This image was produced using the same antibody clone, but in a different formulation, ab8069.

    IHC image showing no staining when ab8069 was used on mouse kidney formalin fixed paraffin embedded tissue, using MOM detection kit, ab127055. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30 mins. The section was incubated with ab8069 at 5 µg/ml for 15 mins at room temperature. DAB was used as the chromogen. The section was counterstained with hematoxylin 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.

  • Flow Cytometry - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Flow Cytometry - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

    This image was produced using the same antibody clone, but in a different formulation, ab8069.

    Overlay histogram showing SV40LT-SMC cells stained with ab8069 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 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 (ab8069, 0.1μg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 0.1μg/1x106 cells) used under the same conditions. Unlabeled sample (blue line) was also used as a control.

    Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

    This antibody gave a positive signal in SV40LT-SMC cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Triton X-100 for 20 min used under the same conditions.

  • Flow Cytometry - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)
    Flow Cytometry - Anti-Vimentin antibody [V9] - BSA and Azide free (ab223871)

    This image was produced using the same antibody clone, but in a different formulation, ab8069.

    Overlay histogram showing MDA-MB-231 cells stained with ab8069 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 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 (ab8069, 1μg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 1μg/1x106 cells) used under the same conditions. Unlabeled sample (blue line) was also used as a control.

    Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

    This antibody gave a positive signal in MDA-MB-231 cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Triton X-100 for 20 min used under the same conditions.

Protocols

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

Datasheets and documents

  • Datasheet download

    Download

References (0)

Publishing research using ab223871? Please let us know so that we can cite the reference in this datasheet.

ab223871 has not yet been referenced specifically in any publications.

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