Recombinant Anti-MMP14 antibody [EP1264Y] - Low endotoxin, Azide free (ab168726)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EP1264Y] to MMP14 - Low endotoxin, Azide free
- Suitable for: ICC/IF, Flow Cyt (Intra), WB, IHC-P, IP
- Knockout validated
- Reacts with: Mouse, Rat, Human
Overview
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Product name
Anti-MMP14 antibody [EP1264Y] - Low endotoxin, Azide free
See all MMP14 primary antibodies -
Description
Rabbit monoclonal [EP1264Y] to MMP14 - Low endotoxin, Azide free -
Host species
Rabbit -
Tested applications
Suitable for: ICC/IF, Flow Cyt (Intra), WB, IHC-P, IPmore details -
Species reactivity
Reacts with: Mouse, Rat, Human
Predicted to work with: Cow -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- IHC-P: Human endometrium carcinoma and kidney tissues. WB: Human fetal spleen, esophagus and lung cancer tissue lysates, mouse and rat spleen tissue lysates; A431 wild-type cell lysate and Caco-2 whole cell lysate. Flow Cyt (intra): HT-1080 cells. IP: A431 cell lysate. ICC/IF: HT-1080 cells and wild-type A431 cells.
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General notes
ab168726 is the carrier-free version of ab194242.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Our Low endotoxin, azide-free formats have low endotoxin level (≤ 1 EU/ml, determined by the LAL assay) and are free from azide, to achieve consistent experimental results in functional assays.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
Storage buffer
Constituent: PBS -
Carrier free
Yes -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EP1264Y -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Conjugation kits
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Isotype control
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KO cell lines
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KO cell lysates
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Recombinant Protein
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab168726 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 |
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ICC/IF |
Use at an assay dependent concentration.
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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 65 kDa.
For unpurified use at 1/2000. |
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IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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IP |
Use at an assay dependent concentration.
For unpurified use at 1/100. |
Notes |
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ICC/IF
Use at an assay dependent concentration. |
Flow Cyt (Intra)
Use at an assay dependent concentration. |
WB
Use at an assay dependent concentration. Predicted molecular weight: 65 kDa. For unpurified use at 1/2000. |
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
IP
Use at an assay dependent concentration. For unpurified use at 1/100. |
Target
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Function
Seems to specifically activate progelatinase A. May thus trigger invasion by tumor cells by activating progelatinase A on the tumor cell surface. May be involved in actin cytoskeleton reorganization by cleaving PTK7. -
Tissue specificity
Expressed in stromal cells of colon, breast, and head and neck. Expressed in lung tumors. -
Sequence similarities
Belongs to the peptidase M10A family.
Contains 4 hemopexin-like domains. -
Domain
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. -
Post-translational
modificationsThe precursor is cleaved by a furin endopeptidase. -
Cellular localization
Membrane. Melanosome. Identified by mass spectrometry in melanosome fractions from stage I to stage IV. - Information by UniProt
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Database links
- Entrez Gene: 281915 Cow
- Entrez Gene: 4323 Human
- Entrez Gene: 17387 Mouse
- Entrez Gene: 81707 Rat
- Omim: 600754 Human
- SwissProt: Q9GLE4 Cow
- SwissProt: P50281 Human
- SwissProt: P53690 Mouse
see all -
Alternative names
- Matrix metallopeptidase 14 (membrane inserted) antibody
- Matrix metalloproteinase 14 antibody
- Matrix metalloproteinase-14 antibody
see all
Images
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All lanes : Anti-MMP14 antibody [EP1264Y] (ab51074) at 1/2000 dilution
Lane 1 : Wild-type A-431 (Human epidermoid carcinoma cell line) whole cell lysate
Lane 2 : MMP14 knockout A-431 (Human epidermoid carcinoma cell line) whole cell lysate
Lane 3 : Caco-2 (Human colorectal adenocarcinoma cell line) whole cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 65 kDa
Observed band size: 54 kDa why is the actual band size different from the predicted?This data was developed using the same antibody clone in a different buffer formulation (ab51074).
Lanes 1 - 3: Merged signal (red and green). Green - ab51074 observed at 54 kDa. Red - loading control, ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.
ab51074 was shown to react with MMP14 in A431 wild-type cells in Western blot. Loss of signal was observed when MMP14 knockout sample was used. A431 wild-type and MMP14 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% Milk in TBS-T (0.1% Tween®) before incubation with ab51074 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
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This data was developed using the same antibody clone in a different buffer formulation (ab51074). ab51074 staining MMP14 in wild-type A431 cells (top panel) and MMP14 knockout A431 cells (bottom panel) (ab261890). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab51074 at 0.2μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8). -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human endometrium carcinoma tissue sections labeling MMP14 with purified ab51074 at 1/100 dilution (1.7 μg/ml). Heat mediated antigen retrieval was performed using EDTA Buffer, PH9. Hematoxylin was used to counterstain. ab97051, a Goat Anti-Rabbit IgG H&L (HRP) secondary antibody was used at 1/500 dilution.
PBS instead of the primary antibody was used as the negative control (inset).
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab51074).
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Ab51074 staining MMP14 in HT-1080 (human fibrosarcoma epithelial cell) cells by ICC/IF (Immunocytochemistry/Immunofluorescence).
Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody at 1/1000 dilution (0.2 μg/ml). An Alexa Fluor® 488 Goat anti-rabbit (ab150077) was used as the secondary antibody at 1/1000 dilution (2 μg/ml). Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594, ab195889) was used as the counterstain antibody at 1/200 dilution (2.5 μg/ml). DAPI was used as a nuclear counterstain. Confocal image showing cytoplasmic and weakly membranous staining in HT-1080 cell line.
Negative control (bottom panels): MCF7 PMID: 19208838.
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ab51074 (purified) at 1/20 dilution (2 µg) immunoprecipitating MMP14 in A431 (human epidermoid carcinoma) whole cell lysate.
Lane 1: A431 whole cell lysate 10ug
Lane 2: ab51074 + A431 whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab51074 in A431 whole cell lysateFor western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.
Blocking and diluting buffer and concentration: 5% NFDM/TBST.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab51074).
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Intracellular Flow Cytometry analysis of MCF7 (Human breast adenocarcinoma epithelial cell, Left) / HT-1080 (Human fibrosarcoma epithelial cell, Right) cells labeling MMP14 with ab51074 at 1/200 dilution (0.1 µg) (red). Goat anti-rabbit IgG (Alexa Fluor®488, ab150077) was used as the secondary antibody at 1/2000 dilution. Rabbit monoclonal IgG (ab172730) / black was used as the isotype control. Cells incubated with secondary antibody only (blue) was used as the unlabeled control. Gated on viable cells.
Positive control (Right panel):HT-1080 cells.
Negative control (Left panel):MCF7 cells.
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ab51074 (unpurified) at 1/500 staining human kidney tissue sections by IHC-P.
The tissue was formaldehyde fixed and a heat mediated antigen retrieval step (in Tris/EDTA) was performed. The tissue was then blocked with serum and incubated with the primary antibody. A biotinylated donkey anti-rabbit IgG was used as the secondary.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab51074).
Protocols
Datasheets and documents
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Datasheet download
References (0)
ab168726 has not yet been referenced specifically in any publications.