Overview

  • Product name
  • Description
    Rabbit polyclonal to Collagen IV
  • Specificity
    ab6586 is designed to bind specifically to NATIVE collagen epitopes composed of multiple subunit strands. Negligible cross-reactivity with Type I, II, III, V or VI collagens. Non-specific cross reaction of anti-collagen antibodies with other human serum proteins or non-collagen extracellular matrix proteins is negligible.
  • Tested applications
    Suitable for: ELISA, IHC-Fr, IP, WB, ICC/IF, IHC-P, IHC-FrFl, IHC-FoFrmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Hamster, Cow, Dog, Human, Pig, Zebrafish, African green monkey, Chinese hamster, Syrian hamster
    Predicted to work with: Mammal
  • Immunogen

    Full length native protein (purified) corresponding to Collagen IV. Collagen Type IV from human and bovine placenta. The immunogen maintains the native conformation of the protein.

  • Positive control
    • human epidermal keratinocytes lysate
  • General notes

    At least 11 genetically distinct gene products are collectively referred to as 'collagen types' or other proteins and proteoglycans of the extracellular matrix. In humans, collagens are composed of about 20 unique protein chains which under go various types of post-translational modifications and are ultimately assembled into a triple helix. This results in great diversity between collagen types. Collagens are highly conserved throughout evolution and are characterized by an uninterrupted "Glycine-X-Y" triplet repeat that is a necessary part of the triple helical structure. For these reasons it is often extremely difficult to generate antibodies with specificities to collagens. The development of type specific antibodies is dependent on NON-DENATURED three-dimensional epitopes. This preparation results in a native conformation of the protein.

     

    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). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
  • Storage buffer
    pH: 8.00
    Preservative: 0.01% Sodium azide
    Constituents: 4.7625% Sodium borate, 0.146% EDTA, 0.435% Sodium chloride
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Purification notes
    Immunoaffinity chromatography using immobilized antigens followed by extensive cross-adsorption against other collagens, human serum proteins and non-collagen extracellular matrix proteins to remove any unwanted specificities.
  • 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 ab6586 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
ELISA 1/5000 - 1/50000.
IHC-Fr 1/50 - 1/200.
IP 1/100.
WB 1/1000 - 1/10000. Use under non reducing condition. This product is not recommended for use under denaturing conditions in WB, IP, and ELISA. We would suggest testing it under native conditions.
ICC/IF Use at an assay dependent concentration. PubMed: 19933193
IF Use at an assay dependent concentration. PubMed: 28153846
IHC-P 1/500. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
IHC-FrFl Use at an assay dependent concentration.
IHC-FoFr Use at an assay dependent concentration.

Target

  • Function
    Type IV collagen is the major structural component of glomerular basement membranes (GBM), forming a 'chicken-wire' meshwork together with laminins, proteoglycans and entactin/nidogen.
    Arresten, comprising the C-terminal NC1 domain, inhibits angiogenesis and tumor formation. The C-terminal half is found to possess the anti-angiogenic activity. Specifically inhibits endothelial cell proliferation, migration and tube formation. Inhibits expression of hypoxia-inducible factor 1alpha and ERK1/2 and p38 MAPK activation. Ligand for alpha1/beta1 integrin.
  • Tissue specificity
    Highly expressed in placenta.
  • Involvement in disease
    Defects in COL4A1 are a cause of brain small vessel disease with hemorrhage (BSVDH) [MIM:607595]. Brain small vessel diseases underlie 20 to 30 percent of ischemic strokes and a larger proportion of intracerebral hemorrhages. Inheritance is autosomal dominant.
    Defects in COL4A1 are the cause of hereditary angiopathy with nephropathy aneurysms and muscle cramps (HANAC) [MIM:611773]. The clinical renal manifestations include hematuria and bilateral large cysts. Histologic analysis revealed complex basement membrane defects in kidney and skin. The systemic angiopathy appears to affect both small vessels and large arteries.
    Defects in COL4A1 are a cause of porencephaly familial (PCEPH) [MIM:175780]. Porencephaly is a term used for any cavitation or cerebrospinal fluid-filled cyst in the brain. Porencephaly type 1 is usually unilateral and results from focal destructive lesions such as fetal vascular occlusion or birth trauma. Type 2, or schizencephalic porencephaly, is usually symmetric and represents a primary defect or arrest in the development of the cerebral ventricles.
  • Sequence similarities
    Belongs to the type IV collagen family.
    Contains 1 collagen IV NC1 (C-terminal non-collagenous) domain.
  • Domain
    Alpha chains of type IV collagen have a non-collagenous domain (NC1) at their C-terminus, frequent interruptions of the G-X-Y repeats in the long central triple-helical domain (which may cause flexibility in the triple helix), and a short N-terminal triple-helical 7S domain.
  • Post-translational
    modifications
    Lysines at the third position of the tripeptide repeating unit (G-X-Y) are hydroxylated in all cases and bind carbohydrates.
    Prolines at the third position of the tripeptide repeating unit (G-X-Y) are hydroxylated in some or all of the chains.
    Type IV collagens contain numerous cysteine residues which are involved in inter- and intramolecular disulfide bonding. 12 of these, located in the NC1 domain, are conserved in all known type IV collagens.
    The trimeric structure of the NC1 domains is stabilized by covalent bonds between Lys and Met residues.
    Proteolytic processing produces the C-terminal NC1 peptide, arresten.
  • Cellular localization
    Secreted > extracellular space > extracellular matrix > basement membrane.
  • Information by UniProt
  • Database links
  • Alternative names
    • Arresten antibody
    • BSVD antibody
    • CO4A1_HUMAN antibody
    • COL4A1 antibody
    • collagen alpha-1(IV) chain antibody
    • collagen type IV alpha 1 chain antibody
    • RATOR antibody
    see all

Anti-Collagen IV antibody images

  • ab6586 staining Collagen IV in Dog liver tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 5% serum for 30 minutes at 25°C; antigen retrieval was by heat mediation in a 10mM citrate buffer, pH 6.0. Samples were incubated with primary antibody (1/200 in PBS + 1x casein) for 1 hour at 37°C. An undiluted HRP-conjugated Horse anti-rabbit IgG polyclonal was used as the secondary antibody.

    See Abreview

  • Immunocytochemical analysis of Mouse 3T3 cell labeling Collagen IV with ab6586 at 1/250 dilution

    See Abreview

  • Anti-Collagen IV antibody (ab6586) at 1/1000 dilution + Baby Hamster Kidney fibroblasts at 100 µg

    Secondary
    HRP-conjugated donkey anti-rabbit polyclonal IgG at 1/4000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Observed band size : 300 kDa (why is the actual band size different from the predicted?)


    Exposure time : 10 seconds

    This image is courtesy of an anonymous Abreview

    Blocked with 5% milk

    See Abreview

  • ab6586 staining rat brain tissue sections (ab4616) by IHC-Fr.  Sections were acetone fixed and blocked with 1% serum for 20 minutes at 25°C.  The primary antibody was diluted 1/300 and incubated with the sample for 30 minutes at 25°C.  A biotinylated goat anti-rabbit IgG antibody was used as the secondary.

     

    See Abreview

  • ab6586 staining Collagen IV in heart tissue by Immunohistochemistry (Frozen sections). The sections were fixed in Acetone prior to blocking with 100% SuperBlock Blocking Buffer for 20 mins at 23°C. The primary antibody was diluted 1/50 and incubated with the sample for 12 hours at 4°C. ab6720 was used as the secondary antibody, diluted 1/100.

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  • ab6586 staining Collagen IV in pig epithelial tissue by Immunohistochemistry (Frozen sections).
    Tissue was fixed in acetone then incubated with ab6586 at a 1/200 dilution for 1 hour at 25°C. The secondary used was ab96886, a goat polyclonal to rabbit IgG - H&L (DyLight® 649), used at a 1/500 dilution.

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  • ab6586 staining Collagen IV in Human corneal epithelial tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde, permeabilized with Triton X-100 and blocked with 10% serum for 1 hour at 20°C; antigen retrieval was enzymatic using trypsin. Samples were incubated with primary antibody (1/500 inPBS + 2%NGS) for 18 hours at 4°C. An Alexa Fluor®488-conjugated Goat anti-rabbit polyclonal (1/500) (ab150077) was used as the secondary antibody.

    See Abreview

  • All lanes : Anti-Collagen IV antibody (ab6586) at 1/1000 dilution

    Lane 1 : Mouse muscle whole tissue lysate
    Lane 2 : Mouse muscle whole tissue lysate

    Lysates/proteins at 50 µg per lane.

    Secondary
    HRP conjugated Goat anti-rabbit IgG polyclonal at 1/5000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Observed band size : 200 kDa (why is the actual band size different from the predicted?)


    Exposure time : 3 minutes

    This image is courtesy of an anonymous Abreview

    Blocked with 5% BSA for 1 hour at 25°C.

    See Abreview

References for Anti-Collagen IV antibody (ab6586)

This product has been referenced in:
  • Moeendarbary E  et al. The soft mechanical signature of glial scars in the central nervous system. Nat Commun 8:14787 (2017). ICC/IF ; Rat . Read more (PubMed: 28317912) »
  • O'Callaghan J  et al. Therapeutic potential of AAV-mediated MMP-3 secretion from corneal endothelium in treating glaucoma. Hum Mol Genet 26:1230-1246 (2017). ICC/IF ; Human . Read more (PubMed: 28158775) »

See all 128 Publications for this product

Product Wall

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (Corneal endothelial cells)
Permeabilization
Yes - Triton 0,3% - 5 minutes
Specification
Corneal endothelial cells
Blocking step
Normal Goat Serum as blocking agent for 18 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 4°C
Fixative
Methanol
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Submitted Aug 21 2017

Application
IHC - Wholemount
Sample
Mouse Tissue (intestine organoids)
Specification
intestine organoids
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Submitted Apr 11 2017

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Tumour plugs)
Antigen retrieval step
Enzymatic - Buffer/Enzyme Used: Trypsin 0.1% in 0.1% CaCl2
Permeabilization
No
Specification
Tumour plugs
Blocking step
BSA (1%) + 10% Goat Serum as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: 25°C
Fixative
Formaldehyde
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Submitted Mar 16 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Chinese hamster Cell lysate - whole cell (Chinese hamster ovary (CHO) cells)
Gel Running Conditions
Reduced Denaturing (4-12% Bolt Bis-Tris Plus gel (MES buffer))
Loading amount
50000 cells
Specification
Chinese hamster ovary (CHO) cells
Blocking step
Li-Cor Odyssey Blocking Buffer (TBS) as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C
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Submitted Dec 31 2016

Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Sample
Rabbit Cell lysate - whole cell (Rabbit Reticulocyte)
Gel Running Conditions
Reduced Denaturing
Loading amount
0.5 µg
Specification
Rabbit Reticulocyte
Blocking step
Li-Cor Odyssey Blocking Buffer (TBS) as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C
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Submitted Nov 21 2016

Application
Immunoprecipitation
Sample
Human Cell lysate - whole cell (A549 (human lung epithelium cells))
Total protein in input
5e+006 cells
Immuno-precipitation step
Other - Protein G Dynabeads
Specification
A549 (human lung epithelium cells)
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Submitted Nov 04 2016

Application
Immunoprecipitation
Sample
Mouse Cell lysate - whole cell (Mouse embryonic fibroblasts (MEFs))
Total protein in input
5e+006 cells
Immuno-precipitation step
Other - Protein G Dynabeads
Specification
Mouse embryonic fibroblasts (MEFs)
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Submitted Nov 04 2016

Application
Immunohistochemistry (Frozen sections)
Sample
Rat Tissue sections (Brain)
Permeabilization
Yes - 0.3 % Triton X-100 included in blocking step
Specification
Brain
Blocking step
10 % Donkey serum, 3 % BSA, 0.3 % triton X-100 in PBS as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 22°C
Fixative
Paraformaldehyde
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Submitted Sep 13 2016

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Sheep Tissue sections (artery)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: 10mM Citrate pH6
Permeabilization
No
Specification
artery
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Fixative
10% normal buffered formalin
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Submitted Sep 01 2016

Application
Western blot
Sample
Hamster Cell lysate - whole cell (BHK-21 (Kidney Fibroblasts))
Gel Running Conditions
Reduced Denaturing (4-12% Bolt Bis-Tris Plus gel)
Loading amount
50000 cells
Specification
BHK-21 (Kidney Fibroblasts)
Blocking step
Li-Cor Odyssey Blocking Buffer (TBS) as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C
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Submitted Jul 22 2016

1-10 of 132 Abreviews or Q&A

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