製品の概要

  • 製品名

  • 製品の詳細

    Rabbit polyclonal to Collagen IV
  • 由来種

    Rabbit
  • 特異性

    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.
  • アプリケーション

    適用あり: ELISA, IHC-Fr, IP, WB, ICC/IF, IHC-P, IHC-FrFl, IHC-FoFrmore details
  • 種交差性

    交差種: Mouse, Rat, Hamster, Cow, Dog, Human, Pig, Zebrafish, African green monkey, Chinese hamster, Syrian hamster
    交差が予測される動物種: Mammals
  • 免疫原

    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.

  • ポジティブ・コントロール

    • Natural Human Collagen IV protein (ab7536) can be used as a positive control in WB. human epidermal keratinocytes lysate
  • 特記事項

    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.

    Abcam recommended secondaries - Goat Anti-Rabbit HRP (ab205718) and Goat Anti-Rabbit Alexa Fluor® 488 (ab150077).

    See other anti-rabbit secondary antibodies that can be used with this antibody.

製品の特性

  • 製品の状態

    Liquid
  • 保存方法

    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.
  • バッファー

    pH: 8.00
    Preservative: 0.01% Sodium azide
    Constituents: 4.7625% Sodium borate, 0.146% EDTA, 0.435% Sodium chloride
  • Concentration information loading...
  • 精製度

    Immunogen affinity purified
  • 特記事項(精製)

    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.
  • 一次抗体 備考

    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.
  • ポリ/モノ

    ポリクローナル
  • アイソタイプ

    IgG
  • 研究分野

アプリケーション

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.

アプリケーション Abreviews 特記事項
ELISA 1/5000 - 1/50000.
IHC-Fr 1/50 - 1/200.
IP 1/100.
WB 1/1000 - 1/10000. Use under non reducing condition. Predicted molecular weight: 160 kDa.

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.

ターゲット情報

  • 機能

    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.
  • 組織特異性

    Highly expressed in placenta.
  • 関連疾患

    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.
  • 配列類似性

    Belongs to the type IV collagen family.
    Contains 1 collagen IV NC1 (C-terminal non-collagenous) domain.
  • ドメイン

    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.
  • 翻訳後修飾

    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.
  • 細胞内局在

    Secreted > extracellular space > extracellular matrix > basement membrane.
  • Information by UniProt
  • 参照データベース

  • 別名

    • Arresten antibody
    • BSVD antibody
    • CO4A1_HUMAN antibody
    • COL4A1 antibody
    • COL4A1 NC1 domain antibody
    • COL4A2 antibody
    • COL4A3 antibody
    • COL4A4 antibody
    • COL4A5 antibody
    • collagen alpha-1(IV) chain antibody
    • Collagen IV Alpha 1 Polypeptide antibody
    • Collagen IV Alpha 2 Polypeptide antibody
    • Collagen Of Basement Membrane Alpha 1 Chain antibody
    • Collagen Of Basement Membrane Alpha 2 Chain antibody
    • Collagen Type IV Alpha 1 antibody
    • collagen type IV alpha 1 chain antibody
    • Collagen Type IV Alpha 2 antibody
    • Collagen Type IV Alpha 3 antibody
    • Collagen Type IV Alpha 4 antibody
    • Collagen Type IV Alpha 5 antibody
    • RATOR antibody
    see all

画像

  • Paraffin-embedded human kidney tissue stained for Collagen IV using ab6586 at 1/400 dilution in immunohistochemical analysis with strong staining observed in glomeruli.

  • Paraffin-embedded human liver tissue stained for Collagen IV using ab6586 at 1/400 dilution in immunohistochemical analysis, strong staining was observed in the sinusoids.

  • 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.

    Predicted band size: 160 kDa
    Observed band size: 300 kDa
    why is the actual band size different from the predicted?


    Exposure time: 10 seconds


    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.

    See Abreview

  • 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.

    See Abreview

参考文献

This product has been referenced in:

  • Zambaiti E  et al. Whole rat stomach decellularisation using a detergent-enzymatic protocol. Pediatr Surg Int 35:21-27 (2019). Read more (PubMed: 30443739) »
  • Gama Sosa MA  et al. Low-level blast exposure disrupts gliovascular and neurovascular connections and induces a chronic vascular pathology in rat brain. Acta Neuropathol Commun 7:6 (2019). Read more (PubMed: 30626447) »
See all 366 Publications for this product

レビューと Q&A

1-10 of 46 Q&A

Question
Answer

I looked at the alignment between the Collagen IV from Hydra vulgaris (http://www.uniprot.org/uniprot/V9GW26) and the immunogen used to raise ab6586, the alignment is only 42%.
Therefore, I would not recommend this antibody for Hydra vulgaris samples.

Read More

Answer

The following antibodies we can recommend as per your requirements. These are tested in IHC-P and WB with mouse samples and are guaranteed.
collagen IV

https://www.abcam.com/collagen-iv-antibody-ab6586.html

anti Laminin

https://www.abcam.com/laminin-antibody-ab128053.html

anti Fibronectin
https://www.abcam.com/fibronectin-antibody-ab2413.html

Read More

Question

I am trying to do fluorescent immunohistochemistry staining for Collagen IV (using ab6586) in paraffin-embedded mouse ovaries. I am trying to use paraffin-embedded mouse kidney as a positive control before staining my mouse ovaries, but have not been able to get consistent positive stain in the kidney. My IHC-P protocol involves: xylene bath x 5 min x 2 Ethanol 100%, 90%, 70% and 30% x 1 min each. 1x PBS wash x 5 min. Heat mediated antigen retrieval with citrate buffered pH 6.0 (I microwave the antigen retrieval soln with the slides in it until boiling (about 2 min) then continue microwaving 15 sec on, 45 sec off, for an additional 10 min) Rinse in 1x PBS x 5 min. 0.2% glycine in PBS x 30 min. Preblock overnight at 4 deg C wtih 10% NGS+10% BSA in PBS. Rinse 1x PBS x 5 min Apply 0.1% sodium borohydride x 10 min Rinse 1x PBS x 5 min Permeabilize with 10% NSG+Saponin (10ul/ml) x 1 hr. Apply primary antibody (1:500 dilution in 10% NGS used) overnight at 4 deg C. Rinse 1x PBS x 5 min. Apply secondary antibody (using AlexaFluor 488-donkey-anti-rabbit at 1:500) and DAPI x 1 hr Rinse 1x PBS x 5 min mount with ScyTek Aqueous mount, seal with nail polish and image. The first time I did this, I saw scant and scattered staining around glomeruli in the renal cortex, but none within them or around tubules. The second time I tried this, I omitted the sodium borohydride and saponin step as it didn't appear others were doing this when I looked at your website, however, that time I saw no staining at all.

Read More
Answer

Thank you for taking time to provide these details. I am sorry to hear that this antibody is not providing satisfactory results.

The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality.

Having reviewed this case, I would like to offer some suggestions to help optimize the results from ab6586. I would also appreciate if you can confirm some further details:



Attempting the described heat-mediated antigen retrieval with a Tris/EDTA ph 9.0 retrieval buffer may be worthwhile. Our recommended buffer formulations and heat-mediated retrieval protocols are given through the link below:

https://www.abcam.com/index.html?pageconfig=resource&rid=11488



It is also worth trying to increase the primary antibody concentration to 1/50 as there is an abReview posted on the webpage that used a 1/50 dilution in mouse tissue in order to visualize COLIV.



Should the suggestions not improve the results, please do let me know.

In the event that a product is not functioning in the species and applications cited on the product datasheet (and the problem has been reported within 6 months of purchase), we would be pleased to provide a free of charge replacement, credit note, or refund.

I hope this information is helpful, and I thank you for your cooperation.

Read More

Answer



With regards to blocking step you can go ahead and incubate with the goat serum and the donkey serum though its not essential but for cleaner results it may be a good idea. You can also use 1% BSA in PBS for blocking.

Primary antibody incubations can be in 1% BSA with both antibodies as a mixture. The same can be done for the secondaries as well.

If you do not want to use BSA then you can use either the goat serum or the rabbit serum to prepare all your antibodies.

Hope this information is useful. Please let me know if I can be of any further assistance.

Read More

Answer

Thank you for taking the time to complete our questionnaire and sending the images. Reviewing this case, I would like to offer some suggestions to help optimise the results from ab6586.

- As a lysis buffer I would suggest using RIPA (radio immuno precipitation assay) buffer due to its higher denaturing level. You can prepare it following the recipe under the next link:

https://www.abcam.com/index.html?pageconfig=resource&rid=11414

- Reduce protein load from 25ug to 15ug.

-For the blocking conditions, instead of using milk, I would optimise the process by using 5% BSA (in TBST) and block for 1-2 hours at room temperature.

-Decrease antibody concentration. The recommended range for WB varies from 1/1000 to 1/10,000. It is worth trying more diluted antibody, and if BSA is used as blocking agent, it is recommended to use the same buffer as antibody dilution buffer.

- In order to dismiss non specific bands coming from the secondary, I would run a non primary control, incubating only with the secondary.

I hope these suggestions will help to improve your results. If our troubleshooting tips do not help, please let us know how you would like to proceed and I would be happy to assist you further in resolving the problem you are experiencing.

No dudes en contactarnos en castellano si lo prefieres.

Read More

Answer

Thank you for contacting us.

Thank you for clarifying that you would prefer to do IHC on paraffin tissue sections rather than frozen tissue sections. For both of these applications I would indeed suggest checking the biocompare website. Ideally you would need a mouse anti-rabbit collagen IV antibody. If this is not possible, then I would try looking for a mouse anti-human collagen IV that has been proven to react with rabbit collagen IV since unfortunately, we do not have this in our catalogue.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Read More

Answer

Thank you for contacting us.

I am sorry to hear you are experiencing difficulties with one of our products. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly.

I will be very happy to help you, and for that, I would appreciate if you could please send me more protocol details. I send you a questionnaire that only take about 5 minutes to be completed, and could help us understand the results obtained and also take the appropriate actions.

Could you also please send the lot number of the vial you have received and from the vial your colleague in Costa Rica did?

All of our products are covered by our Abpromise® guarantee, which ensures that you can trust our products, and they should work in the tested species and applications stated on the datasheet, or we will offer a replacement, credit, or refund, if reported within 6 months of purchase.

To read about our Abpromise policy: https://www.abcam.com/index.html?pageconfig=abpromise

I look forward to receiving your reply. Please do not hesitate to contact us if you need any more advice or information.

Read More

Answer

Thank you for contacting us.

I am sorry, I was under the impression that you wanted to do immunohistochemistry on rabbit tissue. If this is not the case, what application would you like to use the anti-collagen IV antibody in?

With regards to finding an antibody which is likely to work rabbit, I have managed to find the protein sequences of the alpha chain 1,2 and 4 of rabbit collagen IV at:

http://www.uniprot.org/uniprot/Q7M2X2
http://www.uniprot.org/uniprot/Q7M2X5
http://www.uniprot.org/uniprot/P55787

respectively.

However, some of these sequences are incomplete. For the amino acid data we do have, it appears that there is a relatively high % homology between rabbit and human however because, our data is incomplete and it appears that the rabbit protein may be smaller, I am unable to predict whether an antibody is likely to work or not in rabbit.

I would like to recommend checking the Biocompare website which has an excellent antibody search facility that includes many suppliers to see if they have an anti-collagen IV predicted to work in rabbit. The links are:
www.biocompare.com
http://www.biocompare.com/ProductCategories/2045/Antibodies.html

Otherwise, if you would like to test one of the antibodies from our catalogue we have 7 mouse monoclonals which are guaranteed to work in IHC to choose from. If you would like to test one of them, please contact us letting us know which one you would like to test.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Free Rabbit monoclonal antibody with any purchase of a primary antibody, while stocks last! Quote “RABMAB-XBSMG” in your next primary antibody order. For more information, visit the following link: https://www.abcam.com/index.html?pageconfig=resource&rid=15447

Read More

Question
Answer

Thank you for contacting us.

That lot was created in August 2009. We have set an expiration date for that lot for August 2019, so it should still be viable.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Free Rabbit monoclonal antibody with any purchase of a primary antibody, while stocks last! Quote “RABMAB-XBSMG” in your next primary antibody order. For more information, visit the following link: https://www.abcam.com/index.html?pageconfig=resource&rid=15447

Read More

Answer

Thank you for contacting us.

I would recommend not using a rabbit polyclonal on rabbit tissue as it is very likely to give you high background. I have had a look at our catalogue and unfortunately, we do not have any antibodies which are guaranteed to work in rabbit. I am currently contacting our lab to find out whether we have a mouse antibody which is likely to work in rabbit. Do you have the protein sequence of anti-collagen IV for rabbit as this would greatly help determine whether our antibodies could work?

If we have an antibody which is likely to work in rabbit I could offer you a testing discount if you would be willing to test it.

Simply follow these easy steps below to apply for our AbTrial Program:

1. Reply to this email, letting us know you are interested in testing this product.

2. Our scientists will email you an inactive personal discount code for the value of the product.

3. Purchase and test the product at the regular price.

4. Submit your results, including your discount code in the additional notes section of your Abreview.

5. Once the Abreview is submitted, the discount code will become active.

6. Apply your discount code on your next order to receive that value off.

Please let me know if you have any questions about this offer and I would be happy to help you further.

The Terms and Conditions of this offer can be found at: www.abcam.com/abtrial.

Unfortunately, I may not find out from the lab whether our antibodies would be compatible in rabbit by the end of the week.

Regarding Anti-collagen IV, Acris Catalog no. BP5031, was this product bought from us? I have attached our protocol for antigen retrieval.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Free Rabbit monoclonal antibody with any purchase of a primary antibody, while stocks last! Quote “RABMAB-XBSMG” in your next primary antibody order. For more information, visit the following link: https://www.abcam.com/index.html?pageconfig=resource&rid=15447

Read More

1-10 of 46 Q&A

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