製品の概要

  • 製品名
  • 製品の詳細
    Rabbit polyclonal to MMP9
  • 特異性
    The antibody binds to Gelatinase-B, but does not cross react with the other MMP family members (MMP-1, MMP-2, MMP-3). In our hands, we observe a weaker signal in WB in human samples compared to mouse samples (BLAST of full length mouse protein sequence showed 72% homology with the Human MMP9 sequence).
  • アプリケーション
    適用あり: IHC-P, IHC-Fr, WB, IP, ELISA, ICC/IF, ICC, IHC-FoFrmore details
  • 種交差性
    交差種: Mouse, Rat, Dog, Human
  • 免疫原

    Full length protein corresponding to Mouse MMP9.

  • ポジティブ・コントロール
    • HL60 cell lysate. U937 cell lysate. HT1080 cell lysate. Raw 264.7 cell lysate (LPS treated)

アプリケーション

Our Abpromise guarantee covers the use of ab38898 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
IHC-P 1/100 - 1/1000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
IHC-Fr 1/1000. (see Abreview submitted by Greg Gibson)
WB 1/1000. Detects a band of approximately 92 kDa. When using colorimetric substrates such as BCIP/NBT use at a 1/5000 dilution (for chemiluminescent substrates). Detects a band of approximately 92-95 kDa (pro-form) and 82kDa (active form) (Human samples). Mouse MMP9 is larger, and on SDS PAGE gels runs about 102-105 kDa. Dilution optimised using Chromogenic detection.
IP Use at an assay dependent concentration.
ELISA Use at an assay dependent concentration.
ICC/IF 1/500.
ICC Use at an assay dependent concentration.
IHC-FoFr Use at an assay dependent concentration. PubMed: 19295156

ターゲット情報

  • 機能
    May play an essential role in local proteolysis of the extracellular matrix and in leukocyte migration. Could play a role in bone osteoclastic resorption. Cleaves KiSS1 at a Gly-
    -Leu bond. Cleaves type IV and type V collagen into large C-terminal three quarter fragments and shorter N-terminal one quarter fragments. Degrades fibronectin but not laminin or Pz-peptide.
  • 組織特異性
    Produced by normal alveolar macrophages and granulocytes.
  • 関連疾患
    Intervertebral disc disease
    Metaphyseal anadysplasia 2
  • 配列類似性
    Belongs to the peptidase M10A family.
    Contains 3 fibronectin type-II domains.
    Contains 4 hemopexin repeats.
  • ドメイン
    The conserved cysteine present in the cysteine-switch motif binds the catalytic zinc ion, thus inhibiting the enzyme. The dissociation of the cysteine from the zinc ion upon the activation-peptide release activates the enzyme.
  • 翻訳後修飾
    Processing of the precursor yields different active forms of 64, 67 and 82 kDa. Sequentially processing by MMP3 yields the 82 kDa matrix metalloproteinase-9.
    N- and O-glycosylated.
  • 細胞内局在
    Secreted, extracellular space, extracellular matrix.
  • Information by UniProt
  • 参照データベース
  • 別名
    • 82 kDa matrix metalloproteinase-9 antibody
    • 92 kDa gelatinase antibody
    • 92 kDa type IV collagenase antibody
    • CLG 4B antibody
    • CLG4B antibody
    • Collagenase Type 4 beta antibody
    • Collagenase type IV 92 KD antibody
    • EC 3.4.24.35 antibody
    • Gelatinase 92 KD antibody
    • Gelatinase B antibody
    • Gelatinase beta antibody
    • GelatinaseB antibody
    • GELB antibody
    • Macrophage gelatinase antibody
    • MANDP2 antibody
    • Matrix metallopeptidase 9 (gelatinase B, 92kDa gelatinase, 92kDa type IV collagenase) antibody
    • Matrix Metalloproteinase 9 antibody
    • MMP 9 antibody
    • MMP-9 antibody
    • MMP9 antibody
    • MMP9_HUMAN antibody
    • Type V collagenase antibody
    see all

画像

  • All lanes : Anti-MMP9 antibody (ab38898) at 2 µg/ml

    Lane 1 : Natural human MMP9 protein (Proenzyme, monomer) (ab157344)
    Lane 2 : Recombinant Mouse MMP9 protein (ab39309)

    Lysates/proteins at 0.1 µg per lane.

    Secondary
    Infrared labelled goat anti-rabbit (green) at 1/20000 dilution

    Performed under reducing conditions.

    This blot was produced using a 4-12% Bis-Tris gel under the MOPS buffer system. The gel was run at 200V for 60 minutes before being transferred onto a nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour before being incubated with anti-MMP9 antibody (ab38898; 2 microgram per mL)  overnight at 4°C. Antibody binding was detected using infrared labelled goat anti-rabbit (green) antibody (diluted 1:20000) for 1 hour at room temperature before imaging.

  • Anti-MMP9 antibody (ab38898) + Human MMP9

    Observed band size : 88,92 kDa (why is the actual band size different from the predicted?)
    Ab38898 detects a band at 92 Kd (pro-form) and a band at 88 Kd (active form). Mouse MMP9 is slightly larger than human MMP9, and the antibody detects a band at about 105 Kd. It is recommend to concentrate samples by Gelatin-agarose affinity chromatography prior to Western blot usage. A recommended starting concentration for Western blots is 1:1000 when using colorimetric substrates such as BCIP/NBT, and 1:5000 for chemiluminescent substrates. Higher concentration of antibody may be needed for non-human samples.
  • All lanes : Anti-MMP9 antibody (ab38898) at 5 µg/ml

    Lane 1 : Recombinant Human MMP9, His tagged (ab82955) at 0.1 µg
    Lane 2 : U937 whole cell lysate at 100 µg
    Lane 3 : U937 whole cell lysate - treated with PMA and Brefeldin (24 hour treatment) at 100 µg
    Lane 4 : Raw 264.7 (Mouse) whole cell lysate at 100 µg
    Lane 5 : Raw 264.7 (Mouse) whole cell lysate - treated with LPS (6 hour treatment, 1ug/mL) at 100 µg


    Performed under reducing conditions.

    ab38898 detects recombinant Human MMP9 running at ~85 kDa, and endogenous full-length MMP9 in LPS-stimulated cells at ~100 kDa. This antibody also detects a band at 90 kDa in U937 PMA-treated cells. 

     

    ab38898 was incubated at 5 ug/mL and ab8245 (loading control to GAPDH) was diluted at 0.1 ug/mL and both were incubated for 48 hours at 4°C. Blots were developed with goat anti-rabbit IgG (H + L) and goat anti-mouse IgG (H + L) secondary antibodies at 1/10 000 dilution for 1 h at room temperature before imaging.

  • All lanes : Anti-MMP9 antibody (ab38898) at 1/1000 dilution

    Lane 1 : Human lung tissue lysate at 100 µg with 10% Milk for 1 hour at room temperature
    Lane 2 : Human lung tissue lysate at 100 µg with 10% Milk for 1 hour at room temperature
    Lane 3 : MMP-9 KO mice tissue lysates with 10% Milk for 1 hour at room temperature
    Lane 4 : MMP-9 KO mice tissue lysates with 10% Milk for 1 hour at room temperature

    Secondary
    HRP-conjugated donkey anti-rabbit polyclonal at 1/1000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    This image is courtesy of an anonymous Abreview

    Specific observed bands 95-100 kDa

    See Abreview

  • ab38898 staining MMP9 (red) in Mouse Neutrophils and Monocytes cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde and permeabilized with 2%BSA + 0.2% tritonX100 in PBS. Samples were incubated with primary antibody (1/200 in 2%BSA + 0.2% tritonX100 in PBS) for 25 minutes at 23°C. An Alexa Fluor® 568-conjugated Donkey anti-rabbit IgG polyclonal (1/1000) was used as the secondary antibody. DAPI is stained blue

    See Abreview

  • ab38898 staining MMP9 in Mouse Pancreatic carcinoma tissue sections by IHC-P (formaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA for 1 hour at room temperature. Antigen retrieval was by heat mediation in citric acid (pH6). Samples were incubated with primary antibody (1/100) in 1% Aurion BSA for 12 hours. An HRP-conjugated Donkey polyclonal to rabbit IgG (1/100) was used as secondary antibody.

    See Abreview

  • ab38898 staining MMP9 in 6 month-old transgenic zebrafish pancreas (Ihha overexpression) by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections).

    Sections were incubated with primary antibody (1/500) and HRP-conjugated secondary antibody colored using DAB solution. Slides were counterstained with hematoxylin.
  • ab38898 at a 1/1000 dilution staining mouse heart tissue by Immunohistochemistry (Frozen sections). The tissue was removed from a mouse, rinsed in PBS and slowly frozen in supercooled isopentane. 14um sections were made using a cryostat. The sections were acetone fixed and blocked in 2% BSA prior to incubation with the MMP9 antibody. A Cy3 conjugated goat anti-rabbit antibody (ab6939) was used as the secondary antibody. In the image: red staining = MMP9, blue staining = nuclei, green = gelatinase activity.

プロトコール

参考文献

This product has been referenced in:
  • Park JH  et al. Promoting Wound Healing Using Low Molecular Weight Fucoidan in a Full-Thickness Dermal Excision Rat Model. Mar Drugs 15:N/A (2017). IHC-P ; Rat . Read more (PubMed: 28387729) »
  • Chen X  et al. The role of miR-497-5p in myofibroblast differentiation of LR-MSCs and pulmonary fibrogenesis. Sci Rep 7:40958 (2017). WB ; Mouse . Read more (PubMed: 28098218) »

See all 190 Publications for this product

レビューと Q&A

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Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Lymph node)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citrate buffer pH 6
Permeabilization
No
Specification
Lymph node
Blocking step
Dako Protein Block made from Casein X0909 as blocking agent for 10 minute(s) · Concentration: 0.25% · Temperature: 21°C
Fixative
Formaldehyde
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投稿 Jul 18 2017

Application
Western blot
Sample
Pig Cell lysate - whole cell (Pig cell lystate)
Gel Running Conditions
Reduced Denaturing
Loading amount
50 µg
Specification
Pig cell lystate
Blocking step
Milk as blocking agent for 25 minute(s) · Concentration: 5% · Temperature: 23°C
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投稿 Jun 27 2017

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Mammary Tumour Plug)
Antigen retrieval step
Enzymatic - Buffer/Enzyme Used: 0.1% Trypsin in 0.1% CaCl2
Permeabilization
No
Specification
Mammary Tumour Plug
Blocking step
1% BSA + 10% Goat Serum in PBS as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: 25°C
Fixative
Formaldehyde
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投稿 Apr 03 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Rat Tissue lysate - whole (cerebral cortical tissues)
Gel Running Conditions
Reduced Denaturing (10%)
Loading amount
100 µg
Specification
cerebral cortical tissues
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C
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投稿 Apr 03 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Hippocampal astrocyte culture)
Permeabilization
Yes - Methanol
Specification
Hippocampal astrocyte culture
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C
Fixative
Paraformaldehyde
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投稿 Apr 03 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Rat Cell (Rat brain neuronal culture)
Permeabilization
Yes - Methanol
Specification
Rat brain neuronal culture
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C
Fixative
Paraformaldehyde
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投稿 Apr 03 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Sample
Rat Tissue sections (Rat cerebral cortex cryostat section)
Permeabilization
Yes - Methanol
Specification
Rat cerebral cortex cryostat section
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5 · Temperature: 37°C
Fixative
Acetone
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投稿 Apr 03 2017

Application
ELISA
Sample
Mouse Purified protein (Hippocampal Tissues)
Specification
Hippocampal Tissues
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C
Type
Sandwich (Capture)
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投稿 Apr 03 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Sample
Mouse Tissue sections (Mouse brain cryostat sections of hippocampus regio)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Sodium Citrate buffer pH 6.0
Permeabilization
No
Specification
Mouse brain cryostat sections of hippocampus regio
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C
Fixative
Acetone
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投稿 Apr 03 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Tissue lysate - whole (Hippocampal Tissues)
Gel Running Conditions
Reduced Denaturing (10%)
Loading amount
100 µg
Specification
Hippocampal Tissues
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
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投稿 Mar 31 2017

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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