Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-Met (c-Met) antibody [EPR19067] (ab216574)

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Overview

  • Product name

    Anti-Met (c-Met) antibody [EPR19067]
    See all Met (c-Met) primary antibodies

  • Description

    Rabbit monoclonal [EPR19067] to Met (c-Met)

  • Host species

    Rabbit

  • Tested applications

    Suitable for: WB, IHC-P, ICC/IF, Flow Cytmore details

  • Species reactivity

    Reacts with: Human

  • Immunogen

    Recombinant fragment within Human Met (c-Met) aa 1-550. The exact sequence is proprietary.
    Database link: P08581

  • Positive control

    • WB: A549, HeLa and HepG2 whole cell lysates; Human liver lysate; 293T whole cell lysate transfected with a His-tagged human c-Met construct; HeLa whole cell lysate, untreated or treated with PNGase F. IHC-P: Human breast, colon, liver cancer and ovary cancer tissues. ICC/IF: HeLa and A549 cells. Flow Cyt: A549 and HeLa cells.

  • General notes

    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
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

Properties

Applications

Our Abpromise guarantee covers the use of ab216574 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
WB 1/1000. Detects a band of approximately 45-175 kDa (predicted molecular weight: 155 kDa).
IHC-P 1/2000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF 1/1000.
Flow Cyt 1/600.

Target

  • Function

    Receptor for hepatocyte growth factor and scatter factor. Has a tyrosine-protein kinase activity. Functions in cell proliferation, scattering, morphogenesis and survival.

  • Involvement in disease

    Note=Activation of MET after rearrangement with the TPR gene produces an oncogenic protein.
    Note=Defects in MET may be associated with gastric cancer.
    Defects in MET are a cause of hepatocellular carcinoma (HCC) [MIM:114550].
    Defects in MET are a cause of renal cell carcinoma papillary (RCCP) [MIM:605074]. It is a subtype of renal cell carcinoma tending to show a tubulo-papillary architecture formed by numerous, irregular, finger-like projections of connective tissue. Renal cell carcinoma is a heterogeneous group of sporadic or hereditary carcinoma derived from cells of the proximal renal tubular epithelium. It is subclassified into common renal cell carcinoma (clear cell, non-papillary carcinoma), papillary renal cell carcinoma, chromophobe renal cell carcinoma, collecting duct carcinoma with medullary carcinoma of the kidney, and unclassified renal cell carcinoma.
    Note=A common allele in the promoter region of the MET shows genetic association with susceptibility to autism in some families. Functional assays indicate a decrease in MET promoter activity and altered binding of specific transcription factor complexes.
    Note=MET activating mutations may be involved in the development of a highly malignant, metastatic syndrome known as cancer of unknown primary origin (CUP) or primary occult malignancy. Systemic neoplastic spread is generally a late event in cancer progression. However, in some instances, distant dissemination arises at a very early stage, so that metastases reach clinical relevance before primary lesions. Sometimes, the primary lesions cannot be identified in spite of the progresses in the diagnosis of malignancies.

  • Sequence similarities

    Belongs to the protein kinase superfamily. Tyr protein kinase family.
    Contains 3 IPT/TIG domains.
    Contains 1 protein kinase domain.
    Contains 1 Sema domain.

  • Domain

    The kinase domain is involved in SPSB1 binding.

  • Post-translational
    modifications

    Dephosphorylated by PTPRJ at Tyr-1349 and Tyr-1365.

  • Cellular localization

    Membrane.
  • Information by UniProt

  • Database links

    • Alternative names

      • AUTS9 antibody
      • c met antibody
      • D249 antibody
      • Hepatocyte growth factor receptor antibody
      • HGF antibody
      • HGF receptor antibody
      • HGF/SF receptor antibody
      • HGFR antibody
      • MET antibody
      • Met proto oncogene antibody
      • Met proto oncogene tyrosine kinase antibody
      • MET proto oncogene, receptor tyrosine kinase antibody
      • Met proto-oncogene (hepatocyte growth factor receptor) antibody
      • Met proto-oncogene antibody
      • Met protooncogene antibody
      • MET_HUMAN antibody
      • Oncogene MET antibody
      • Par4 antibody
      • Proto-oncogene c-Met antibody
      • RCCP2 antibody
      • Scatter factor receptor antibody
      • SF receptor antibody
      • Tyrosine-protein kinase Met antibody
      see all

    Images

    • Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Lane 1: Hap1 wildtype cell lysate (20 µg)
      Lane 2: MET Hap1 knockout cell lysate (20 µg)
      Lane 3: HeLa wildtype cell lysate (20 µg)
      Lane 4: MET HeLa knockout cell lysate (20 µg)

      Lanes 1 - 4: Merged signal (red and green). Green - ab51067 observed at 160 kDa. Red - loading control, ab8245 observed at 37 kDa.

      ab51067 was shown to react with Met (c-Met) in HeLa wildtype. Loss of signal was observed when knockout sample ab256991 was used. Wild-type and Met (c-Met) knockout samples were subjected to SDS-PAGE. ab51067 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

    • Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Lane 1: Wild-type HAP1 whole cell lysate (40 µg)
      Lane 2: MET knockout HAP1 whole cell lysate (40 µg)

      Lanes 1 - 2: Merged signal (red and green). Green - ab216574 observed at 155 kDa. Red - loading control, ab18058, observed at 130 kDa.

      ab216574 was shown to specifically recognize MET in wild-type HAP1 cells along with additional cross reactive bands. No bands were observed when MET knockout samples were used. Wild-type and MET knockout samples were subjected to SDS-PAGE. ab216574 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/10,000 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 1/10,000 dilution for 1hr at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Immunohistochemical analysis of paraffin-embedded human breast tissue labeling Met (c-Met) with ab216574 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use.

      Membranous staining on human breast is observed.

      Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunocytochemistry/ Immunofluorescence - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Immunocytochemistry/ Immunofluorescence - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Met (c-Met) with ab216574 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

      Confocal image showing cytoplasmic staining on HeLa cell line.

      The nuclear counterstain is DAPI (blue).

      Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)) at 1/200 dilution (red).

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) at 1/1000 dilution.

    • Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      All lanes : Anti-Met (c-Met) antibody [EPR19067] (ab216574) at 1/1000 dilution

      Lane 1 : A549 (Human lung carcinoma cell line) whole cell lysate
      Lane 2 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
      Lane 3 : HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

      Predicted band size: 155 kDa
      Observed band size: 45-175 kDa
      why is the actual band size different from the predicted?


      Exposure time: 30 seconds


      Blocking/Dilution buffer: 5% NFDM/TBST.

      This antibody detects Pro-c-Met (175 kDa), c-Met beta subunit (145 kDa) [PMID: 22418436], c-Met alpha subunit (45 kDa) [PMID: 8710887] and a cleavage c-Met fragment (85 kDa) [PMID: 11786517].

    • Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Anti-Met (c-Met) antibody [EPR19067] (ab216574) at 1/1000 dilution + Human liver lysate at 10 µg

      Secondary
      Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/10000 dilution

      Predicted band size: 155 kDa
      Observed band size: 100-150 kDa why is the actual band size different from the predicted?


      Exposure time: 30 seconds


      Blocking/Dilution buffer: 5% NFDM/TBST.

      In human liver the antibody detected c-Met beta subunit (145 kDa) [PMID: 22418436] and a cleavage c-Met fragment (100 kDa) [PMID: 18187039].

    • Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      All lanes : Anti-Met (c-Met) antibody [EPR19067] (ab216574) at 1/10000 dilution

      Lane 1 : 293T whole cell lysate (Human epithelial cell line from embryonic kidney) transfected with an empty expression vector
      Lane 2 : 293T whole cell lysate transfected with a His-tagged human c-Met construct

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

      Predicted band size: 155 kDa
      Observed band size: 150-175 kDa why is the actual band size different from the predicted?


      Exposure time: 1 second


      Blocking and Diluting buffer and concentration: 5% NFDM /TBST 

    • Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Western blot - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      All lanes : Anti-Met (c-Met) antibody [EPR19067] (ab216574) at 1/1000 dilution

      Lane 1 : Untreated HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
      Lane 2 : HeLa whole cell lysate treated with PNGase F

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

      Predicted band size: 155 kDa


      Exposure time: 1 minute


      Blocking/Dilution buffer: 5% NFDM/TBST.

      After partial deglycosylation, the band of c-Met beta subunit (145 kDa) is shifted to 125 kDa. The 85 kDa glycosylated band is reduced and a ~60 kDa band appears.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Immunohistochemical analysis of paraffin-embedded human colon tissue labeling Met (c-Met) with ab216574 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use.

      Membranous staining on human colon is observed.

      Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Immunohistochemical analysis of paraffin-embedded human liver cancer tissue labeling Met (c-Met) with ab216574 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use.

      Membranous staining on tumor cells of human liver cancer is observed.

      Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Immunohistochemical analysis of paraffin-embedded human ovary cancer tissue labeling Met (c-Met) with ab216574 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use.

      Cytoplasmic and membranous staining on tumor cells of human ovary cancer is observed.

      Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunocytochemistry/ Immunofluorescence - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Immunocytochemistry/ Immunofluorescence - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A549 (Human lung carcinoma cell line) cells labeling Met (c-Met) with ab216574 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

      Confocal image showing cytoplasmic staining on A549 cell line.

      The nuclear counterstain is DAPI (blue).

      Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)) at 1/200 dilution (red).

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) at 1/1000 dilution.

    • Flow Cytometry - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Flow Cytometry - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Flow cytometric analysis of 4% paraformaldehyde-fixed A549 (Human lung carcinoma cell line) cells labeling Met (c-Met) with ab216574 at 1/600 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

    • Flow Cytometry - Anti-Met (c-Met) antibody [EPR19067] (ab216574)
      Flow Cytometry - Anti-Met (c-Met) antibody [EPR19067] (ab216574)

      Flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Met (c-Met) with ab216574 at 1/600 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

    References

    This product has been referenced in:

    • Shi W  et al. MACC-1 antibody target therapy suppresses growth and migration of non-small cell lung cancer. Mol Med Rep 16:7329-7336 (2017). Read more (PubMed: 28944826) »
    See 1 Publication for this product

    Publishing research using ab216574? Please let us know so that we can cite the reference in this datasheet.

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