Overview

  • Product name

    Anti-HGF antibody [EPR12230]
    See all HGF primary antibodies
  • Description

    Rabbit monoclonal [EPR12230] to HGF
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, IPmore details
    Unsuitable for: IHC-P
  • Species reactivity

    Reacts with: Human
    Does not react with: Mouse, Rat
  • Immunogen

    Synthetic peptide within Human HGF aa 100-200 (Cysteine residue). The exact sequence is proprietary.
    Database link: P14210

  • Positive control

    • Human placenta and U-87 MG cell lysates and Human HGF (aa 32-728) recombinant protein. IP: Human placenta lysate.
  • General notes

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

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab178395 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 1/200. Predicted molecular weight: 83 kDa.

For unpurified use at 1/1000- 1/5000.

IP 1/10 - 1/100.
  • Application notes
    Is unsuitable for IHC-P.
  • Target

    • Function

      Potent mitogen for mature parenchymal hepatocyte cells, seems to be a hepatotrophic factor, and acts as a growth factor for a broad spectrum of tissues and cell types. Activating ligand for the receptor tyrosine kinase MET by binding to it and promoting its dimerization.
    • Involvement in disease

      Deafness, autosomal recessive, 39
    • Sequence similarities

      Belongs to the peptidase S1 family. Plasminogen subfamily.
      Contains 4 kringle domains.
      Contains 1 PAN domain.
      Contains 1 peptidase S1 domain.
    • Information by UniProt
    • Database links

    • Alternative names

      • DFNB39 antibody
      • F TCF antibody
      • Fibroblast derived tumor cytotoxic factor antibody
      • Hepatocyte growth factor (hepapoietin A; scatter factor) antibody
      • Hepatocyte growth factor antibody
      • Hepatocyte growth factor beta chain antibody
      • Hepatocyte growth factor precursor antibody
      • Hepatopoietin A antibody
      • Hepatopoietin-A antibody
      • Hgf antibody
      • HGF_HUMAN antibody
      • HGFB antibody
      • HPTA antibody
      • Lung fibroblast derived mitogen antibody
      • OTTHUMP00000161349 antibody
      • OTTHUMP00000206710 antibody
      • OTTHUMP00000206711 antibody
      • OTTHUMP00000206712 antibody
      • OTTHUMP00000206713 antibody
      • OTTHUMP00000206730 antibody
      • Scatter factor antibody
      • SF antibody
      see all

    Images

    • Anti-HGF antibody [EPR12230] (ab178395) at 1/1000 dilution (Purified) + U-87 MG (Human glioblastoma-astrocytoma epithelial cell) whole cell lysates at 20 µg

      Secondary
      Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

      Predicted band size: 83 kDa
      Observed band size: 83 kDa

    • Anti-HGF antibody [EPR12230] (ab178395) at 1/200 dilution (Purified) + Human placenta lysates at 15 µg

      Secondary
      Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

      Predicted band size: 83 kDa
      Observed band size: 83 kDa

    • ab178395 (purified) at 1:30 dilution (2µg) immunoprecipitating HGF in Human placenta lysate.
      Lane 1 (input): Human placenta lysate 10µg
      Lane 2 (+): ab178395 & Human placenta lysate
      Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab178395 in Human placenta lysate
      For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1:1000 dilution.
      Blocking and diluting buffer: 5% NFDM/TBST.

    References

    This product has been referenced in:

    • Deying W  et al. CAF-derived HGF promotes cell proliferation and drug resistance by up-regulating the c-Met/PI3K/Akt and GRP78 signalling in ovarian cancer cells. Biosci Rep 37:N/A (2017). Read more (PubMed: 28258248) »
    • Kwon MJ  et al. Programmed death ligand-1 and MET co-expression is a poor prognostic factor in gastric cancers after resection. Oncotarget 8:82399-82414 (2017). Read more (PubMed: 29137273) »
    See all 3 Publications for this product

    Customer reviews and Q&As

    1-2 of 2 Abreviews or Q&A

    Answer

    Thank you for contacting us.

    The western blot image shown on the datasheet for ab178395 was obtained using denaturing and reducing conditions.

    I have copied below the WB protocols that have been used. Please note these would be a guideline only and may require individual optimization.

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




    Cell Lysate preparation



    Harvest 3.0*108 cells.
    Add 3 ml 1×Hot lysis buffer .
    Mix the cells and buffer well, then boil the tube for 15-30 min. (Take the tube out and mix them between whiles during incubation. )
    Sonication. 3 seconds, intervals 3 seconds, 25-30 times (40kW) until the cell clumps all scattered and the liquid is clear.
    Centrifugation. Centrifugate the tube at 10,000rpm at 4℃ for 10min and then put the supernatant at 4℃ temporarily.
    Protein concentration quantitative analysis by BCA method.
    Adjust concentration of the cell lysate to 4 mg/ml with 1×Hot lysis buffer.
    Adjust to 2mg/ml protein with 2×sample buffer, keep 5min in boiling water. Then ready to use.





    1×Hot lysis Buffer

    10 mM Tris-Hcl(pH8.0)

    1%SDS

    1.0 mM Na-Orthovanadate

    ddH2O



    2×Sample Buffer

    125mM Tris-Hcl(pH6.8)

    2.5% SDS

    0.04% Bromophenol Blue

    25% Glycerol:

    713mM ß-Mercaptoethanol:

    ddH2O











    Tissue Lysate preparation



    Cut the frozen tissue into small piece with scissor. Grind the small piece into powder in mortar and transfer them to pre-cold centrifuge tube.



    * The scissors, mortar and pestle should be pre-cold in -80 degree overnight.

    Add 1×Hot lysis buffer, 2 times the volume of the tissue power.
    Mix the tissue and buffer well, then incubate tube at 95-100°C for 15-30 min. (Take the tube out and mix them between whiles during incubation. )
    Sonication. 3 seconds, intervals 3 seconds, 25-30 times (40kW) until the tissue clumps all scattered and the liquid is clear.
    Centrifugation. Centrifugate the tube at 10,000rpm at 4℃ for 10min and then put the supernatant at 4℃ temporarily.
    Measure the protein concentration by BCA method.
    Adjust concentration of the tissue lysate to 4 mg/ml with 1×Hot lysis buffer.
    Adjust to 2mg/ml protein with 2×sample buffer, keep 5min in boiling water. Then ready to use.





    * 1×Hot lysis buffer:

    10 mM Tris-Hcl(pH8.0)

    1%SDS

    1.0 mM Na-Orthovanadate

    ddH2O





    *2×Sample Buffer:

    125mM Tris-Hcl(pH6.8)

    2.5% SDS

    0.04% Bromophenol Blue

    25% Glycerol:

    713mM ß- Mercaptoethanol:

    ddH2O









    Western blot protocol



    1. Take the membrane out of 4℃ refrigerator. Treat the membrane with 99.5% methanol for 15 sec..

    2. Rinse the membrane with deionized water for 5 min.

    3. Block the membrane using 5% NFDM/TBST. Swing slightly for overnight at 4℃ on shaking table.

    4. Wash the membrane with 1×TBST for 10 min.

    5. Incubate the membrane with suitable dilution of primary antibody in 5% NFDM/TBST for 1h at RT on shaking table. Swing slightly.

    6. Wash the membrane 3 times with 1×TBST, 10 min per time.

    7. Incubate the membrane with 1:1000 dilution of the secondary antibody in 5% NFDM/TBST for 1h at RT on shaking table. Swing slightly.

    *Secondary antibody information: Pierce, Goat Anti-Rabbit IgG, (H+L), Peroxidase Conjugated.

    8. Wash the membrane with 1×TBST 3 times,10min per time.

    9. Incubate the membrane with ECL substrate for 5min. at RT. (Mix reagent 1 and reagent 2 at the ratio of 1:1. Before using, place the mixture in dark place for 5min.)

    10. Remove excess ECL substrate and sandwich the membrane in transparent plastic wrap.

    11. Acquire image using darkroom development techniques. Expose the membrane for 3 min. (If the signal is very strong, re-expose the membrane for less minute).

    Read More

    Answer

    When ab178395 was validated in flow, permeabilized A549 cells were used.

    Read More

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