Overview

  • Product name
    Anti-Fibronectin antibody (Biotin)
    See all Fibronectin primary antibodies
  • Description
    Rabbit polyclonal to Fibronectin (Biotin)
  • Host species
    Rabbit
  • Conjugation
    Biotin
  • Specificity
    negligible cross-reactivity with Type I, II, III, IV, V or VI Collagens or Laminin. Non-specific cross reaction of anti-Fibronectin antibodies with other human serum proteins or non-Fibronectin extracellular matrix proteins is negligible.
  • Tested applications
    Suitable for: ICC/IF, ELISA, IHC-P, IP, WBmore details
  • Species reactivity
    Reacts with: Mouse, Cow, Human
    Predicted to work with: Mammals
  • Immunogen

    Fibronectin was purified from Human plasma by binding to a denatured gelatin column followed by elution with high concentrations of arginine. The eluted material was further purified by gel filtration. Immunization occurred after single-band purity was assessed by SDS-PAGE

  • General notes


    This antibody is well suited to detect extracellular matrix proteins in normal as well as disease state tissues. Disruption of tissue organization is the hallmark of neoplasia. Malignant lesions can be distinguished from benign by examining the breakdown of basement membranes and loss of 3-dimensional architecture. Malignant cells are presumed to use matrix metalloproteases to degrade barriers created by the extracellular matrix which then allows metastasis to occur. Collagenases, stomelysins and gelatinases can collectively degrade all of the various components of the extracellular matrix, including fibrillar and non-fibrillar collagens and basement membrane glycoproteins.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage buffer
    pH: 7.2
    Preservative: 0.01% Sodium azide
    Constituents: 0.878% Sodium chloride, 1% BSA, 0.424% Potassium phosphate
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Purification notes
    This product has been prepared by immunoaffinity chromatography using immobilized antigens followed by extensive cross-adsorption against human serum proteins and collagen and non-collagen extracellular matrix proteins to remove any unwanted specificities. Typically less than 1% cross reactivity against other extracellular matrix proteins was detected by ELISA against purified standards.
  • Primary antibody notes
    This antibody is well suited to detect extracellular matrix proteins in normal as well as disease state tissues. Disruption of tissue organization is the hallmark of neoplasia. Malignant lesions can be distinguished from benign by examining the breakdown of basement membranes and loss of 3-dimensional architecture. Malignant cells are presumed to use matrix metalloproteases to degrade barriers created by the extracellular matrix which then allows metastasis to occur. Collagenases, stomelysins and gelatinases can collectively degrade all of the various components of the extracellular matrix, including fibrillar and non-fibrillar collagens and basement membrane glycoproteins.
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab6584 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
ICC/IF Use at an assay dependent concentration. PubMed: 20686611
ELISA 1/4000 - 1/8000.
IHC-P 1/50 - 1/200.
IP Use at an assay dependent concentration.
WB 1/5000 - 1/10000.

Target

  • Function
    Fibronectins bind cell surfaces and various compounds including collagen, fibrin, heparin, DNA, and actin. Fibronectins are involved in cell adhesion, cell motility, opsonization, wound healing, and maintenance of cell shape. Involved in osteoblast compaction through the fibronectin fibrillogenesis cell-mediated matrix assembly process, essential for osteoblast mineralization. Participates in the regulation of type I collagen deposition by osteoblasts.
    Anastellin binds fibronectin and induces fibril formation. This fibronectin polymer, named superfibronectin, exhibits enhanced adhesive properties. Both anastellin and superfibronectin inhibit tumor growth, angiogenesis and metastasis. Anastellin activates p38 MAPK and inhibits lysophospholipid signaling.
  • Tissue specificity
    Plasma FN (soluble dimeric form) is secreted by hepatocytes. Cellular FN (dimeric or cross-linked multimeric forms), made by fibroblasts, epithelial and other cell types, is deposited as fibrils in the extracellular matrix. Ugl-Y1, Ugl-Y2 and Ugl-Y3 are found in urine.
  • Involvement in disease
    Glomerulopathy with fibronectin deposits 2
  • Sequence similarities
    Contains 12 fibronectin type-I domains.
    Contains 2 fibronectin type-II domains.
    Contains 16 fibronectin type-III domains.
  • Developmental stage
    Ugl-Y1, Ugl-Y2 and Ugl-Y3 are present in the urine from 0 to 17 years of age.
  • Post-translational
    modifications
    Sulfated.
    It is not known whether both or only one of Thr-2064 and Thr-2065 are/is glycosylated.
    Forms covalent cross-links mediated by a transglutaminase, such as F13A or TGM2, between a glutamine and the epsilon-amino group of a lysine residue, forming homopolymers and heteropolymers (e.g. fibrinogen-fibronectin, collagen-fibronectin heteropolymers).
    Phosphorylated by FAM20C in the extracellular medium.
    Proteolytic processing produces the C-terminal NC1 peptide, anastellin.
  • Cellular localization
    Secreted, extracellular space, extracellular matrix.
  • Information by UniProt
  • Database links
  • Alternative names
    • CIG antibody
    • Cold insoluble globulin antibody
    • Cold-insoluble globulin antibody
    • DKFZp686F10164 antibody
    • DKFZp686H0342 antibody
    • DKFZp686I1370 antibody
    • DKFZp686O13149 antibody
    • ED B antibody
    • Fibronectin 1 antibody
    • FINC antibody
    • FINC_HUMAN antibody
    • FN antibody
    • FN1 antibody
    • FNZ antibody
    • GFND antibody
    • GFND2 antibody
    • LETS antibody
    • Migration stimulating factor antibody
    • MSF antibody
    • Ugl-Y3 antibody
    see all

Images

  • Immunofluorescence analysis of U87-MG cells staining Fibronectin (red) with ab6584.
    Cells were treated with 10-6M Dexamethasone, before blocking and incubating with primary antibody for 1 hour at room temperature. An Alexa Fluor® 568-conjugated anti-rabbit IgG was used as the secondary antibody.

  • Human Aortic Endothelial Cells grown in MCDB 131+ SmGM-2 media were seeded on a PET membrane. After one week, the cells were blocked with PBS containing 0.1% BSA for 15 min. The membrane was incubated with primary antibody (ab6584) for 30 min  and then with secondary antibody Alexa Fluor® 488 for 1 hr.

    This picture was kindly supplied as part of the review submitted by Gary Nackman and Ildiko Entersz.

  • Immunohistochemical analysis of formalin-fixed, paraffin-embedded human kidney tissue sections at pH 6.0, labelling Fibronectin with ab6584 at a concentration of 10 µg/mL for 1 hour at room temperature. The secondary used was a rabbit peroxidase conjugate at a 1/10,000 dilution, incubated for 45 minutes at room temperature. Fibronectin is shown as a brown hue against a purple counterstain labelling nuclear DNA. The negative control on right (B) is without the primary antibody. Magnifications are as shown.

  • Immunohistochemical analysis of formalin-fixed, paraffin-embedded human kidney tissue sections at pH 9.0, labelling Fibronectin with ab6584 at a concentration of 10 µg/mL for 1 hour at room temperature. The secondary used was a rabbit peroxidase conjugate at a 1/10,000 dilution, incubated for 45 minutes at room temperature. Fibronectin is shown as a brown hue against a purple counterstain labelling nuclear DNA. The negative control on right (B) is without the primary antibody. Magnifications are as shown.

References

This product has been referenced in:
  • Nair M  et al. Dexamethasone-Mediated Upregulation of Calreticulin Inhibits Primary Human Glioblastoma Dispersal Ex Vivo. Int J Mol Sci 19:N/A (2018). Read more (PubMed: 29443896) »
  • Raghunathan VK  et al. Glaucomatous cell derived matrices differentially modulate non-glaucomatous trabecular meshwork cellular behavior. Acta Biomater 71:444-459 (2018). ICC/IF . Read more (PubMed: 29524673) »
See all 24 Publications for this product

Customer reviews and Q&As

1-10 of 10 Abreviews or Q&A

Answer

Thank you for your enquiry about our fibronectin antibodies. The antibody ab6584 is biotin conjugated and has been successfully used in IF (ICC) by a user of the antibody who left a review which can be accessed via the online datasheet. The reviewer did not leave many details about the cells or protocol used so you may wish to contact him by clicking on his name on the sheet. As the antibody is biotin conjugated you will need to add an extra-avidin fluorescence conjugate (or avidin), so that the avidin part will bind to the biotin molecules (so you won't need a secondary antibody per se). You can purchase extra-avidin FITC for example and incubate it 1-3 hours with the cells; we currently have no information about the use of ab6584 in IF on sections. We have a similar anti fibronectin antibody unconjugated, ab299, but it has not been tested in IF/ICC or IHC, therefore we do not know if it will work in these applications. Please do not hesitate to contact me if you would like some protocol help with our antibodies or any further information,

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Answer

For testing 1ug of antigen, the minimum of antibody used is 1:8000. The maximum is 1:4000. We don't have other data to show the sensitivity. If you have any additional questions, please contact us again.

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Question
Answer

1:4000 to 1:8000 dilution can detect 1.0ug of Fibronectin. If you have any additional questions, please contact us again.

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Answer

I have not found reports of staining of fibronectin in the cytoplasm of plasma cells, however it is known that plasma cells adhere to fibronectin (the fibronectin inducing not only the adherence and spread of plasma cells but also their migration). I therefore think that the staining you are observing is not specific (especially using normal rabbit serum as blocking agent) and would like to suggest the following changes to your protocol: -what type of IHC are you testing, is it paraffin embedded tissue? If so you need to consider antigen retrieval methods (heat mediated, enzymatic) - you did not mention quenching the endogenous peroxidases prior to start the staining protocol, please make sure you do as the DAB step will give high background if you have not incubated the tissue in H2O2 at the beginning of the protocol to quench endogenous peroxidase -block the tissue in normal goat/horse serum (not rabbit, as the primary antibody will bind to the serum, giving high background/non specific staining) -try decreasing the concentration of the primary antibody and changing the incubation time, temperature and the presence of triton in the dilution buffer -Incubation in ABC could be reduced to 1/2h I hope these recommendations will help you, please do not hesitate to contact me again if you still have problems,

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Answer

This antibody does not require antigen retrieval but it may increase staining. We normally recommend the pressure cooker method of antigen retrieval.

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Question
Answer

450 kDa non-reduced 225 kDa fully reduced

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Answer

This antibody will cross-ract with mouse fibronectin using formalin fixed paraffin embedded tissues.

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Answer

Yes, this antibody is available in an unconjugated form, ab299

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Question
Answer

The extracellular matrix proteins are well conserved, particularly between mammals, so human will cross-react with pig.

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