Overview

  • Product name
    Anti-Src antibody [EPR5496]
    See all Src primary antibodies
  • Description
    Rabbit monoclonal [EPR5496] to Src
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WB, IHC-Pmore details
    Unsuitable for: Flow Cyt or IP
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human Src aa 1-100. The exact sequence is proprietary.
    Database link: P12931

  • Positive control
    • Hela, 293T, SH-SY5Y and HUVEC cell lysates; Human kidney tissue and Human colonic adenocarcinoma tissue; HeLa cells
  • General notes

    A trial size is available to purchase for this antibody.

     

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

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab109381 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 1/10000 - 1/50000. Predicted molecular weight: 60 kDa.
IHC-P 1/250 - 1/500. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
  • Application notes
    Is unsuitable for Flow Cyt or IP.
  • Target

    • Function
      Non-receptor protein tyrosine kinase that plays pivotal roles in numerous cellular processes such as proliferation, migration, and transformation. In concert with PTK2B, plays an important role in osteoclastic bone resorption. Both the formation of a SRC-PTK2B complex, and SRC kinase activity are necessary for this function. Once it is recruited to the activated integrins, by PTK2B, it phosphorylates CBL which in turn induces the activation and recruitment of phosphatidylinositol 3-kinase to the cell membrane in a signaling pathway that is critical for osteoclast function. Promotes energy production in osteoclasts by activating mitochondrial cytochrome C oxidase. Phosphorylates RUNX3 and COX2 on tyrosine residues, TNK2 on 'Tyr-284' and CBL on 'Tyr-731'. Enhances DDX58/RIG-I-elicited antiviral signaling.
    • Sequence similarities
      Belongs to the protein kinase superfamily. Tyr protein kinase family. SRC subfamily.
      Contains 1 protein kinase domain.
      Contains 1 SH2 domain.
      Contains 1 SH3 domain.
    • Post-translational
      modifications
      Dephosphorylated at Tyr-530 by PTPRJ (By similarity). Phosphorylated on Tyr-530 by c-Src kinase (CSK). The phosphorylated form is termed pp60c-src. Dephosphorylated by PTPRJ at Tyr-419. Normally maintained in an inactive conformation with the SH2 domain engaged with Tyr-530, the SH3 domain engaged with the SH2-kinase linker, and Tyr-419 dephosphorylated. Dephosphorylation of Tyr-530 as a result of protein tyrosine phosphatase (PTP) action disrupts the intramolecular interaction between the SH2 domain and Tyr-530, Tyr-419 can then become autophosphorylated, resulting in SRC activation. Phosphorylation of Tyr-530 by CSK allows this interaction to reform, resulting in SRC inactivation.
      S-nitrosylation is important for activation of its kinase activity.
    • Cellular localization
      Cell membrane. Mitochondrion inner membrane.
    • Information by UniProt
    • Database links
    • Form
      This protein is known to be similar in amino acid sequence to HCK (P08631), LCK (P06239), FYN (P06241), YES1 (P07947), and LYN (P07948). Therefore, cross-reactivity with these homologous proteins may be observed. We would be happy to provide immunogen alignment information upon request.
    • Alternative names
      • ASV antibody
      • Avian sarcoma virus antibody
      • c SRC antibody
      • CDNA FLJ14219 fis clone NT2RP3003800 highly similar to Rattus norvegicus tyrosine protein kinase pp60 c src mRNA antibody
      • cSrc antibody
      • EC 2.7.10.2 antibody
      • Neuronal CSRC tyrosine specific protein kinase antibody
      • Neuronal SRC antibody
      • Oncogene SRC antibody
      • OTTHUMP00000174476 antibody
      • OTTHUMP00000174477 antibody
      • p60 Src antibody
      • p60-Src antibody
      • p60Src antibody
      • pp60c src antibody
      • pp60c-src antibody
      • pp60csrc antibody
      • Proto oncogene tyrosine protein kinase Src antibody
      • Proto-oncogene c-Src antibody
      • Proto-oncogene tyrosine-protein kinase Src antibody
      • Protooncogene SRC antibody
      • Protooncogene SRC Rous sarcoma antibody
      • Src antibody
      • SRC Oncogene antibody
      • SRC proto oncogene non receptor tyrosine kinase antibody
      • SRC_HUMAN antibody
      • SRC1 antibody
      • Tyrosine kinase pp60c src antibody
      • Tyrosine protein kinase SRC 1 antibody
      • Tyrosine protein kinase SRC1 antibody
      • v src avian sarcoma (Schmidt Ruppin A2) viral oncogene homolog antibody
      • V src sarcoma (Schmidt Ruppin A 2) viral oncogene homolog (avian) antibody
      • v src sarcoma (Schmidt Ruppin A 2) viral oncogene homolog avian antibody
      see all

    Images

    • Lanes 1, 5 and 9: Wild-type HAP1 cell lysate (20 µg)
      Lanes 2, 6 and 10: Src knockout HAP1 cell lysate (20 µg)
      Lanes 3, 7 and 11: HeLa cell lysate (20 µg)
      Lanes 4, 8 and 12: A431 cell lysate (20 µg)
      Lanes 1, 2, 3 and 4: Green signal from target – ab109381 observed at 60 kDa
      Lanes 5, 6, 7 and 8: Red signal from loading control – ab8245 observed at 37 kDa
      Lanes 9, 10, 11 and 12: Merged (red and green) signal

      ab109381 was shown to specifically react with Src when Src knockout samples were used. Wild-type and Src knockout samples were subjected to SDS-PAGE. ab109381 and ab8245 (loading control to GAPDH) were diluted 1/10 000 and 1/2000 respectively and incubated overnight at 4ºC. 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 1/10 000 dilution for 1 h at room temperature before imaging.

    • All lanes : Anti-Src antibody [EPR5496] (ab109381) at 1/10000 dilution

      Lane 1 : Hela cell lysate
      Lane 2 : 293T cell lysate
      Lane 3 : SH-SY5Y cell lysate
      Lane 4 : HUVEC cell lysate

      Lysates/proteins at 10 µg per lane.

      Predicted band size: 60 kDa

    • ab109381, at 1/250 dilution, staining Src in paraffin-embedded Human kidney tissue by Immunohistochemistry.
    • Lane 1: Wild-type HAP1 cell lysate (20 µg)
      Lane 2: Src knockout HAP1 cell lysate (20 µg)
      Lane 3: HeLa cell lysate (20 µg)
      Lane 4: A431 cell lysate (20 µg)

       

      Lanes 1 - 4: Merged signal (red and green). Green - ab109381 observed at 60 kDa. Red - loading control, ab8245, observed at 37 kDa.

      This western blot image is a comparison between ab109381 and a competitor's top cited rabbit polyclonal antibody.

    • ab109381, at 1/250 dilution, staining Src in paraffin-embedded Human colonic adenocarcinoma tissue by Immunohistochemistry.

    References

    This product has been referenced in:
    • Socodato R  et al. Redox tuning of Ca2+ signaling in microglia drives glutamate release during hypoxia. Free Radic Biol Med 118:137-149 (2018). Mouse . Read more (PubMed: 29501565) »
    • Dandoulaki M  et al. Src activation by Chk1 promotes actin patch formation and prevents chromatin bridge breakage in cytokinesis. J Cell Biol 217:3071-3089 (2018). WB . Read more (PubMed: 29954829) »
    See all 10 Publications for this product

    Customer reviews and Q&As

    1-4 of 4 Abreviews or Q&A

    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (colon carcinoma cells)
    Gel Running Conditions
    Non-reduced Denaturing
    Loading amount
    50 µg
    Specification
    colon carcinoma cells
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

    Abcam user community

    Verified customer

    Submitted Sep 06 2018

    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (U2OS)
    Gel Running Conditions
    Reduced Denaturing
    Loading amount
    30 µg
    Specification
    U2OS
    Blocking step
    Milk as blocking agent for 3 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C

    Abcam user community

    Verified customer

    Submitted May 25 2018

    Answer

    Thank you for contacting us.

    There is nothing we can do about this problem. The problem could be with HRP conjugate of secondary antibody or even the reagents. ECL sometime gives faint staining then the other high sensitive developing reagents e.g. ECL+; this is very common with antibodies. I would suggest trying ECL+ or together high intensity developing reagents.

    I hope this information will be helpful. Should you have any question please do not hesitate to contact me.

    Read More

    Question

    Product code: 109381
    Lot number: GR48347-2
    Inquiry: Here is questionair filled by our customer in Russia: 3) Description of the problem: faint signal in ECL using antibody dilution 1 : 1000 (in passport 1 : 10000 – 1 : 50000 and "Super Signal West Dura Extended Duration Substrate" (Thermo Scientific) 4) Sample preparation: Type of sample (whole cell lysates, fraction, recombinant protein…): whole cell lysates Lysis buffer: 50 mM Тris-HCl, pH 7.5, 150 mM NaCl, 1% Triton X-100 Protease inhibitors: 1 mM PMSF, complete protease inhibitor cocktail tablet (Roche) Phosphatase inhibitors: 1 mM o-vanadate, 50 mM NaF, 2 mM ЕDTA Reducing agent: 5% β-mercaptoethanol Boiling for ≥5 min? yes/no: yes Protein loaded ug/lane or cells/lane: 50 ug Positive control: lysate of HEK293 cells transiently transfected with plasmid encoding wt c-Src, lysate of NIH 3T3 cells Negative control: WB w/o 1-st Abs 5) Percentage of gel: 10% Type of membrane: nitrocellulose Protein transfer verified: Poncean staining Blocking agent and concentration: 5% dry skim milk Blocking time: 1,5 h Blocking temperature: -4oC 6) Primary antibody (If more than one was used, describe in “additional notes”) : only one Concentration or dilution: 1 : 1000 (in passport 1 : 10000 – 1 : 50000 Diluent buffer: PBST, 1% dry skim milk Incubation time: overnight Incubation temperature: -4oC 7) Secondary antibody: anti-rabbit IgG (whole molecule) HRP, Sigma, Product number A6154 Species: Produced in Goat Reacts against: rabbit monoclonal Concentration or dilution: 1 : 5000 Diluent buffer: PBST, 1% dry skim milk Incubation time: 2 h Incubation temperature: -4oC Fluorochrome or enzyme conjugate: enzyme conjugate 8) Washing after primary and secondary antibodies: Buffer: PBST Number of washes: 4 times 9) Detection method 10) How many times have you run this staining? For last 2 years Do you obtain the same results every time? - Yes What steps have you altered to try and optimize the use of this antibody? According to above mentioned protocol

    Read More
    Answer

    Thank you for your email. I am sorry to hear that you are experiencing problems using this antibody.

    I have few more questions to ask in order to understand the problem;

    - Could you provide image of the staining?
    - Though we have used this antibody at 1/10,000 in our own lab however the final dilution depends on the experiment and varies from lab to lab even with lysates. I would suggest trying 1/500 dilution to see if the result improves.

    I will look forward to receiving the images.

    Read More

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

    Sign up