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Anti-TIMP3 抗体 - Carboxyterminal end (ab39185)

Reviews (1)Q&A (5)References (4)
Western blot - Anti-TIMP3 antibody - Carboxyterminal end (ab39185)
  • Immunocytochemistry/ Immunofluorescence - Anti-TIMP3 antibody - Carboxyterminal end (ab39185)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TIMP3 antibody - Carboxyterminal end (ab39185)

Key features and details

  • Rabbit polyclonal to TIMP3 - Carboxyterminal end
  • Suitable for: WB, ICC/IF, IHC-P
  • Reacts with: Human
  • Isotype: IgG

製品の概要

  • 製品名

    Anti-TIMP3 antibody - Carboxyterminal end
    TIMP3 一次抗体 製品一覧

  • 製品の詳細

    Rabbit polyclonal to TIMP3 - Carboxyterminal end

  • 由来種

    Rabbit

  • 特異性

    This antibody recognizes both the glycosylated and unglycosylated forms of TIMP3, and works against native or reduced TIMP3. It does not cross react with the other TIMP family members (TIMP1, TIMP2, TIMP4).

  • アプリケーション

    適用あり: WB, ICC/IF, IHC-Pmore details

  • 種交差性

    交差種: Human

  • 免疫原

    Synthetic peptide based on the carboxyterminal region of Human TIMP3.

  • ポジティブ･コントロール

    • Human amd mouse TIMP3.

  • 特記事項

    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

製品の特性

アプリケーション

The Abpromise guarantee

Abpromise保証は、 次のテスト済みアプリケーションにおけるab39185の使用に適用されます

アプリケーションノートには、推奨の開始希釈率がありますが、適切な希釈率につきましてはご検討ください。

アプリケーション Abreviews 特記事項
WB
1/1000 - 1/5000. Detects a band of approximately 21 kDa (predicted molecular weight: 24 kDa). Used under non reducing conditions.
ICC/IF
Use a concentration of 1 µg/ml.
IHC-P (1)
Use at an assay dependent concentration.
特記事項
WB
1/1000 - 1/5000. Detects a band of approximately 21 kDa (predicted molecular weight: 24 kDa). Used under non reducing conditions.
ICC/IF
Use a concentration of 1 µg/ml.
IHC-P
Use at an assay dependent concentration.

ターゲット情報

  • 機能

    Complexes with metalloproteinases (such as collagenases) and irreversibly inactivates them by binding to their catalytic zinc cofactor. May form part of a tissue-specific acute response to remodeling stimuli. Known to act on MMP-1, MMP-2, MMP-3, MMP-7, MMP-9, MMP-13, MMP-14 and MMP-15.

  • 関連疾患

    Defects in TIMP3 are the cause of Sorsby fundus dystrophy (SFD) [MIM:136900]. SFD is a rare autosomal dominant macular disorder with an age of onset in the fourth decade. It is characterized by loss of central vision from subretinal neovascularization and atrophy of the ocular tissues. Generally, macular disciform degeneration develops in the patients eye within 6 months to 6 years.

  • 配列類似性

    Belongs to the protease inhibitor I35 (TIMP) family.
    Contains 1 NTR domain.

  • 細胞内局在

    Secreted > extracellular space > extracellular matrix.
  • Information by UniProt

  • 参照データベース

    • 別名

      • HSMRK222 antibody
      • K222 antibody
      • K222TA2 antibody
      • Metalloproteinase inhibitor 3 antibody
      • MIG 5 protein antibody
      • MIG5 protein antibody
      • Protein MIG 5 antibody
      • Protein MIG-5 antibody
      • SFD antibody
      • Sorsby fundus dystrophy pseudoinflammatory antibody
      • TIMP 3 antibody
      • TIMP metallopeptidase inhibitor 3 antibody
      • TIMP-3 antibody
      • TIMP3 antibody
      • TIMP3_HUMAN antibody
      • Tissue Inhibitor of Metalloproteinase 3 antibody
      • Tissue inhibitor of metalloproteinases 3 antibody
      • Tissue inhibitor of metalloproteinases3 antibody
      see all

    画像

    • Western blot - Anti-TIMP3 antibody - Carboxyterminal end (ab39185)
      Western blot - Anti-TIMP3 antibody - Carboxyterminal end (ab39185)
      Anti-TIMP3 antibody - Carboxyterminal end (ab39185) at 1 µg/ml + Lung (Human) Tissue Lysate at 10 µg

      Secondary
      Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

      Developed using the ECL technique.

      Performed under reducing conditions.

      Predicted band size: 24 kDa
      Observed band size: 24 kDa
      Additional bands at: 100 kDa. We are unsure as to the identity of these extra bands.


      Exposure time: 90 seconds
    • Immunocytochemistry/ Immunofluorescence - Anti-TIMP3 antibody - Carboxyterminal end (ab39185)
      Immunocytochemistry/ Immunofluorescence - Anti-TIMP3 antibody - Carboxyterminal end (ab39185)
      ICC/IF image of ab39185 stained HeLa cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab39185, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TIMP3 antibody - Carboxyterminal end (ab39185)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TIMP3 antibody - Carboxyterminal end (ab39185)
      Ab39185 staining Human normal placenta. Staining is localized to the cytoplasm or excreted.
      Left panel: with primary antibody at 2 ug/ml. Right panel: isotype control.
      Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 AR buffer citrate EDTA pH 9.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS), then incubated with primary antibody for 20 minutes, and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.

    参考文献 (4)

    ab39185 を使用した論文を発表された方は、こちらまでお知らせください。データシートに掲載させていただきます。

    ab39185 は 4 報の論文で使用されています。

    • Niu T  et al. Long non-coding RNA RPSAP52 upregulates Timp3 by serving as the endogenous sponge of microRNA-365 in diabetic retinopathy. Exp Ther Med 20:246 (2020). PubMed: 33178344
    • Pricola Fehnel K  et al. Using urinary bFGF and TIMP3 levels to predict the presence of juvenile pilocytic astrocytoma and establish a distinct biomarker signature. J Neurosurg Pediatr 18:396-407 (2016). PubMed: 27314542
    • Fiorentino L  et al. Loss of TIMP3 underlies diabetic nephropathy via FoxO1/STAT1 interplay. EMBO Mol Med 5:441-55 (2013). Mouse . PubMed: 23401241
    • Thomay AA  et al. Disruption of interleukin-1 signaling improves the quality of wound healing. Am J Pathol 174:2129-36 (2009). WB ; Mouse . PubMed: 19389930

    レビューと Q&A

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    1-6 of 6 Abreviews or Q&A

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-TIMP3 antibody - Carboxyterminal end

     Excellent
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Blocking step
    Casein in PBS as blocking agent for 10 minute(s) · Concentration: 0.25% · Temperature: 21°C
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citrate Buffer pH 6
    Sample
    Human Tissue sections (Kidney)
    Specification
    Kidney
    Permeabilization
    No
    Fixative
    Formaldehyde
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Dr. Francesca Conserva

    Verified customer

    投稿 Jul 12 2013

    Question

    immunogen sequence or range

    Read More
    Answer

    Thank you for contacting us.
    I have checked, but unfortunately, for these antibodies we do not have more immunogen information than what is on the datasheet. The source where we receive these 3 antibodies from, considers any immunogen information proprietary and therefore we do not have this information ourselves.
    I'm very sorry about this. I know this will be very disappointing for you, but I wish I could be of more help.
    Please do not hesitate to contact us if you need any more advice or information.
    Use our products? Submit an Abreview. Earn rewards!
    https://www.abcam.com/abreviews

    Read More

    Question

    I'm using ab39185 in WB to detect the protein in mouse retina lysate and only seeing a band around 30 kDa, and nothing at the expected 24 kDa.

    Read More
    Answer

    Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

    I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement with ab39184.

    To check the status of the order please contact our Customer Service team and reference this number.

    Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.

    I wish you the best of luck with your research.

    Read More

    Question

    LOT NUMBER Gr 8727-1 ORDER NUMBER -- NOT SPECIFIED -- DESCRIPTION OF THE PROBLEM I'm obtaining bands above 30 kDa for ab39184 and ab39185 (TIMP antibodies). In the description of the antibodies ab39184 and ab39185 is: Anti-TIMP3 antibody - Carboxyterminal end (ab39185) WB: 1/1000 - 1/5000. Used under non reducing conditions. Detects a band of approximately 21 kDa (predicted molecular weight: 24 kDa). Anti-TIMP3 antibody - Loop 1 (ab39184) WB: 1/1000 - 1/5000. Dilution optimised using Chromogenic detection. Detects a band of approximately 24 , 30 kDa (predicted molecular weight: 24 kDa). My bands are above 30 KDa, what should I do? SAMPLE Type of sample: cell lysates of mouse PRIMARY ANTIBODY Primary antibody (If more than one was used, describe in “additional notes”): only TIMP3 (ab 39184) Concentration or dilution: 1:200 Diluent buffer: TTBS 1x (TBS 10x and distilled water) Incubation time: overnight Incubation temperature: 8ºC DETECTION METHOD ECL ANTIBODY STORAGE CONDITIONS as recommended on the datasheet. SAMPLE PREPARATION Lysis buffer: RIPA Boiling for ≥5 min? yes AMOUNT OF PROTEIN LOADED Protein loaded ug/lane or cells/lane: 30 ug/ protein ELECTROPHORESIS/GEL CONDITIONS Non reducing gel. Percentage of gel: gradient 5-20% TRANSFER AND BLOCKING CONDITIONS Type of membrane: nitrocellulose membrane Protein transfer verified: yes Blocking agent and concentration: milk 5% Blocking time: 1 hour Blocking temperature: room temperature SECONDARY ANTIBODY Secondary antibody: anti-rabbit IgG HRP conjugated ( Amersham NA934) Species: Reacts against: rabbit Concentration or dilution: 1:5000 Diluent buffer: TTBS 1x Incubation time: 1h Incubation temperature: room temperature Fluorochrome or enzyme conjugate: HRP Washing after primary and secondary antibodies: Buffer: TTBS Number of washes: primary, 3x (5 min.) secondary, 3x (15 min.) DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? Concentration of antibody

    Read More
    Answer

    Thank you for taking the time to contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from these antibodies. The details you have kindly provided will enable us to investigate this case for you and also gives us vital information for our monitoring of product quality. 1) When testing our antibodies, our lab uses 5% BSA as a blocking reagent, so I recommend switching to this instead of milk. For unknown reasons some antibodies give stronger, more specific signals on blots blocked with BSA instead of milk, so doing this may improve the results you are seeing, and reduce the non-specific bands. An example of the above is the western blot obtained with the Abcam GAPDH antibody ab9385 : Click here for the western blot image using ab9385 (or use the following: www.abcam.com/ab9385).   2) I am unsure if the bands you are observing are around 80 kDa, as the resolution of the higher molecular weight marker bands is suboptimal. In case no bands have run out of the gel and you count from the bottom, the TIMP3 bands could also be around 60 kDa. If that is the case they may represent multimers of the protein (32 kDa dimers were reported according to the literature) or complexes with metalloproteinases. Therefore, I would recommend boiling the samples for more than 5 minutes (e.g. 10min) which may help to reduce disulfide bridges and break up complex structures. We are happy to offer this technical support. In the event that the product is not functioning in the applications cited on the product data sheet (and the problem has been reported within 6 months of purchase), we will be pleased to provide a credit note, free of charge replacement or refund. I hope this information is helpful, thank you for your cooperation. Should the suggestions not improve the results, please do not hesitate to contact me again with the further requested details.  

    Read More

    Question

    Hello, Thanks for your reply. Attached is a file of my blot that was probed with the anti-TIMP3, ab39185. In this blot we included some conditioned medium from melanoma cells that we know have TIMP3 protein (proteomic data). We did probe this blot with anti-TIMP 1 purchased from other company and the band was at the correct molecular weight. So we know the OVC extracts are OK. Your suggestion that the 50kDa band may be a TIMP3 dimer, is acceptable but I have not came across any reference of its existence. If you know of any references, please pass along :). Would it be possible for you to send a small aliquot of other anti-TIMP3 ab that has been used by other researchers in Western blotting for us to confirm if indeed the 50kDa band is a TIMP3 dimer.

    Read More
    Answer

    Thank you for providing the additional information and western blot data. It appears that ab39185 detects a protein of the expected molecular weight in the conditioned medium however the 50kDa band is seen in the OVC tissue extracts. And the band in the OVC samples seems to be quite specific. I have found TIMP3 can form dimers but, as I've found, only regarding the following: "Mutations to TIMP-3, all of which introduce an extra cysteine residue into exon 5 (which forms part of the COOH-terminal domain of the molecule), have been linked to the macular degenerative disease Sorsby’s fundus dystrophy" in: The Journal of Biological Chemistry, 273, 16778-16781. Also we do not have free samples available.

    Read More

    Question

    Hi, I recently purchased anti-TIMP3 antibody (AB39185) from your company for Western blot. I am detecting a very strong band at 50 KDa in ovarian cancer tissue samples. A very weak band of the predicted size for this ab, 20 kDa could be seen only with a long exposure. What can you suggest what could be the reason of this strong band?

    Read More
    Answer

    Thank you for your enquiry. Just a few additional questions regarding your western blot: 1. Have you used any other antibodies for western blotting with this ovarian cancer tissue extract? If so, what were the results? 2. The 50kDa band you see may be a TIMP3 dimer. Have you proped any other human or mouse tissue or cell lysates with ab39185 as a comparison? 3. It might be helpful to view your results. If you could, please forward along an image of your western blot. Thanks very much for the additional information.

    Read More

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