Recombinant
RabMAb

Recombinant Anti-Vimentin (phospho S56) antibody [EPR21084] (ab217673)

Overview

  • Product name

    Anti-Vimentin (phospho S56) antibody [EPR21084]
    See all Vimentin primary antibodies
  • Description

    Rabbit monoclonal [EPR21084] to Vimentin (phospho S56)
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, Dot blot, ICC/IF, IP, Flow Cytmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human Vimentin aa 1-100 (phospho S56). The exact sequence is proprietary.
    Database link: P08670

  • Positive control

    • WB: HeLa treated with 100 ng/ml nocodazole (ab120630) for 18 hours, whole cell lysate. ICC/IF: HeLa cells. IP: HeLa treated with 100 ng/ml nocodazole (ab120630) for 18 hours, whole cell lysate.
  • General notes

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

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab217673 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
WB 1/1000. Detects a band of approximately 57 kDa (predicted molecular weight: 54 kDa).
Dot blot 1/1000.
ICC/IF 1/1000.
IP 1/30.
Flow Cyt 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.
  • Information by UniProt
  • Database links

  • 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

  • All lanes : Anti-Vimentin (phospho S56) antibody [EPR21084] (ab217673) at 1/1000 dilution

    Lane 1 : Untreated HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate
    Lane 2 : HeLa (human epithelial cell line from cervix adenocarcinoma) treated with 100 ng/ml nocodazole (ab120630) for 18 hours, whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Developed using the ECL technique.

    Predicted band size: 54 kDa
    Observed band size: 57 kDa
    why is the actual band size different from the predicted?


    Exposure time: 23 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Ab217673 staining Vimentin (phospho S56) in HeLa (human cervix adenocarcinoma epithelial cell) cells by Flow Cytometry. Cells were fixed using 80% Methanol and permeabilized with 0.1% Tween-20. The sample was incubated with primary antibody at 1:500 dilution. An Alexa Fluor® 488 Goat anti-rabbit IgG (ab150077) was used as a secondary antibody. Rabbit monoclonal IgG (ab172730) was used as an isotype control (left). Cells were pre-treated with 20 μg/ml RNase A for 30 minutes to eliminate the non-specific binding between RNA and PI (Propidium iodide). Vimentiin (phospho S56) is highly expressed in mitotic cells. (PMID: 16260496)

  • Dot blot analysis of Vimentin (phospho S56) labeled with ab217673 at 1/1000 dilution.

    Lane 1: Vimentin (phospho S56) peptide.
    Lane 2: Vimentin non-phospho peptide.

    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution was used as secondary antibody.

    Blocking and dilution buffer: 5% NFDM/TBST.

    Exposure time: 1 minute.

    The blot was developed on a BIO-RAD® ChemiDoc™ MP instrument.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling Vimentin (phospho S56) with ab217673 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing strong positive staining in HeLa cells in M phase.

    The nuclear counterstain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) at 1/200 dilution (red).

    Secondary antibody only control: PBS instead of primary antibody, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

  • Vimentin (phospho S56) was immunoprecipitated from 0.35 mg of HeLa (human epithelial cell line from cervix adenocarcinoma) treated with 100 ng/ml nocodazole (ab120630) for 18 hours, whole cell lysate with ab217673 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab217673 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

    Lane 1: HeLa treated with 100 ng/ml nocodazole (ab120630) for 18 hours, whole cell lysate 10 μg (Input).
    Lane 2: ab217673 IP in HeLa treated with 100 ng/ml nocodazole (ab120630) for 18 hours, whole cell lysate.
    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab217673 in HeLa treated with 100 ng/ml nocodazole (ab120630) for 18 hours, whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

References

ab217673 has not yet been referenced specifically in any publications.

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