Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)

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Western blot - Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)
  • Immunocytochemistry/ Immunofluorescence - Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)
  • Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [Y531] to Nck - BSA and Azide free
  • Suitable for: ICC/IF, WB, IHC-P, Flow Cyt
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Overview

  • Product name

    Anti-Nck antibody [Y531] - BSA and Azide free
    See all Nck primary antibodies

  • Description

    Rabbit monoclonal [Y531] to Nck - BSA and Azide free

  • Host species

    Rabbit

  • Specificity

    ab235389 recognises Nck.

  • Tested applications

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

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Synthetic peptide within Human Nck aa 200-300. The exact sequence is proprietary. Close to the SH2 domain.

  • Positive control

    • WB: Hela and NIH/3T3 cell lysates. ICC/IF: HeLa cells. IHC-P: Human breast carcinoma tissue.

  • General notes

    Ab235389 is the carrier-free version of ab32120. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    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.

    ab235389 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    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
    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.

    Reproducibility is key to advancing scientific discovery and accelerating scientists’ next breakthrough.

    Abcam is leading the way with our range of recombinant antibodies, knockout-validated antibodies and knockout cell lines, all of which support improved reproducibility.

    We are also planning to innovate the way in which we present recommended applications and species on our product datasheets, so that only applications & species that have been tested in our own labs, our suppliers or by selected trusted collaborators are covered by our Abpromise™ guarantee.

    In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.

    We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.

    Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.

    Please check that this product meets your needs before purchasing. If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, as well as customer reviews and Q&As.

Properties

Applications

Our Abpromise guarantee covers the use of ab235389 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
ICC/IF Use at an assay dependent concentration.
WB Use at an assay dependent concentration. Detects a band of approximately 47 kDa.
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Flow Cyt Use at an assay dependent concentration.

Target

  • Function

    Adapter protein which associates with tyrosine-phosphorylated growth factor receptors or their cellular substrates. Maintains low levels of EIF2S1 phosphorylation by promoting its dephosphorylation by PP1. Plays a role in the DNA damage response, not in the detection of the damage by ATM/ATR, but for efficient activation of downstream effectors, such as that of CHEK2.

  • Sequence similarities

    Contains 1 SH2 domain.
    Contains 3 SH3 domains.

  • Post-translational
    modifications

    Phosphorylated on Ser and Tyr residues.

  • Cellular localization

    Cytoplasm. Endoplasmic reticulum. Nucleus. Mostly cytoplasmic, but shuttles between the cytoplasm and the nucleus. Import into the nucleus requires the interaction with SOCS7. Predominantly nuclear following genotoxic stresses, such as UV irradiation, hydroxyurea or mitomycin C treatments.
  • Information by UniProt

  • Database links

    see all

  • Alternative names

    • Cytoplasmic protein NCK1 antibody
    • Melanoma Nck protein antibody
    • MGC12668 antibody
    • NCK 1 antibody
    • NCK adaptor protein 1 antibody
    • NCK alpha antibody
    • NCK antibody
    • NCK tyrosine kinase antibody
    • Nck-1 antibody
    • NCK1 antibody
    • NCK1_HUMAN antibody
    • NCKalpha antibody
    • Non catalytic region of tyrosine kinase antibody
    • SH2/SH3 adaptor protein NCK alpha antibody
    • SH2/SH3 adaptor protein NCK-alpha antibody
    see all

Images

  • Western blot - Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)
    Western blot - Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)
    All lanes : Anti-Nck antibody [Y531] (ab32120) at 1/1000 dilution

    Lane 1 : Wild-type HeLa cell lysate
    Lane 2 : NCK1 knockout HeLa cell lysate
    Lane 3 : NIH/3T3 cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution

    Observed band size: 50 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 (ab32120).

    Lanes 1-3: Merged signal (red and green). Green - ab32120 observed at 50 kDa. Red - loading control ab8245 observed at 36 kDa. 

     ab32120 Anti-Nck antibody [Y531] was shown to specifically react with Nck in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265323 (knockout cell lysate ab258059) was used.  Wild-type and Nck knockout samples were subjected to SDS-PAGE.  ab32120 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2. 5 hours 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.

  • Immunocytochemistry/ Immunofluorescence - Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)
    Immunocytochemistry/ Immunofluorescence - Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)

    Immunocytochemistry/ Immunofluorescence analysis of HeLa (human cervix adenocarcinoma epithelial cell) cells labeling Nck with purified ab32120 at 1/250 dilution (8 µg/mL). Cells were fixed in 100% Methanol.Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/mL) dilution. DAPI (blue) was used as nuclear counterstain.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32120).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)

    This IHC data was generated using the same anti-Nck antibody clone, Y531, in a different buffer formulation (cat# ab32120).

    ab32120 (1/100), staining human breast carcinoma by immunohistochemistry, Paraffin embedded tissue.

    Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

  • Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)
    Anti-Nck antibody [Y531] - BSA and Azide free (ab235389)

Protocols

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

References (0)

Publishing research using ab235389? Please let us know so that we can cite the reference in this datasheet.

ab235389 has not yet been referenced specifically in any publications.

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