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Anti-AHNAK antibody [EM-09] (ab68556)

Reviews (1)Q&A (4)References (3)
Western blot - Anti-AHNAK antibody [EM-09] (ab68556)
  • Immunocytochemistry - Anti-AHNAK antibody [EM-09] (ab68556)
  • Immunocytochemistry - Anti-AHNAK antibody [EM-09] (ab68556)
  • Immunohistochemistry (Frozen sections) - Anti-AHNAK antibody [EM-09] (ab68556)
  • Flow Cytometry - Anti-AHNAK antibody [EM-09] (ab68556)

Key features and details

  • Mouse monoclonal [EM-09] to AHNAK
  • Suitable for: WB, Flow Cyt, IHC-Fr, ICC
  • Reacts with: Mouse, Human
  • Isotype: IgG1

Overview

  • Product name

    Anti-AHNAK antibody [EM-09]
    See all AHNAK primary antibodies

  • Description

    Mouse monoclonal [EM-09] to AHNAK

  • Host species

    Mouse

  • Tested applications

    Suitable for: WB, Flow Cyt, IHC-Fr, ICCmore details

  • Species reactivity

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

  • Immunogen

    Recombinant fragment corresponding to Human AHNAK (N terminal).
    Database link: Q09666

  • Positive control

    • WB: 1% lauryl maltoside lysate of HeLa cells. ICC/IF: Human primary fibroblasts. HeLa cells. IHC-Fr: Mouse tongue tissue. Flow Cyt: HeLa cells.

  • General notes

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. 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, along with publications, customer reviews and Q&As

Properties

  • Form

    Liquid

  • Storage instructions

    Shipped at 4°C. Upon delivery aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.

  • Storage buffer

    pH: 7.4
    Preservative: 0.097% Sodium azide
    Constituent: PBS
  • Concentration information loading...

  • Purity

    Protein A purified

  • Purification notes

    ab68556 is purified from hybridoma culture supernatant. Purity >95% by SDS-PAGE.

  • Clonality

    Monoclonal

  • Clone number

    EM-09

  • Isotype

    IgG1

  • Research areas

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab68556 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
Use at an assay dependent concentration. Predicted molecular weight: 629 kDa.
Flow Cyt
Use 1µg for 106 cells.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
IHC-Fr
Use at an assay dependent concentration.
ICC
1/250.

Permeabilization is required.
Notes
WB
Use at an assay dependent concentration. Predicted molecular weight: 629 kDa.
Flow Cyt
Use 1µg for 106 cells.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
IHC-Fr
Use at an assay dependent concentration.
ICC
1/250.

Permeabilization is required.

Target

  • Function

    May be required for neuronal cell differentiation.

  • Sequence similarities

    Contains 1 PDZ (DHR) domain.

  • Cellular localization

    Nucleus.
  • Information by UniProt

  • Database links

    • Alternative names

      • AHNAK antibody
      • AHNAK nucleoprotein (desmoyokin) antibody
      • AHNAK nucleoprotein antibody
      • AHNAKRS antibody
      • AHNK_HUMAN antibody
      • Desmoyokin antibody
      • Fragments antibody
      • MGC5395 antibody
      • Neuroblast differentiation associated protein AHNAK antibody
      • Neuroblast differentiation-associated protein AHNAK antibody
      • PM227 antibody
      see all

    Images

    • Western blot - Anti-AHNAK antibody [EM-09] (ab68556)
      Western blot - Anti-AHNAK antibody [EM-09] (ab68556)
      Anti-AHNAK antibody [EM-09] (ab68556) at 2 µg/ml
      Performed under non-reducing conditions.

      Predicted band size: 629 kDa



      The material was 1% lauryl maltoside lysate of HeLa cells, non-reducing conditions, immunoprecipitated with another anti-AHNAK1 antibody. 4% stacking gel, 6% separating gel, U=190 V, 60 min. Blotting: 30 mA, 90 min. Detection antibody concentration: 2 microgram/ml. Please note the conditions were not optimalized.
    • Immunocytochemistry - Anti-AHNAK antibody [EM-09] (ab68556)
      Immunocytochemistry - Anti-AHNAK antibody [EM-09] (ab68556)
      Human primary fibroblasts stained with ab68556 (green). Actin filaments were decorated by phalloidin (red) and cell nuclei stained with DAPI (blue).
    • Immunocytochemistry - Anti-AHNAK antibody [EM-09] (ab68556)
      Immunocytochemistry - Anti-AHNAK antibody [EM-09] (ab68556)

      ab68556 at 1/250 dilution staining AHNAK (red) in HeLa cells by Immunocytochemistry/Immunofluorescence. Nuclei were stained with DAPI (blue).

    • Immunohistochemistry (Frozen sections) - Anti-AHNAK antibody [EM-09] (ab68556)
      Immunohistochemistry (Frozen sections) - Anti-AHNAK antibody [EM-09] (ab68556)
      Ab68556 (red) staining AHNAK in murine tongue by immunohistochemistry using frozen tissue sections. Actin filaments were decorated by phalloidin (green), cell nuclei stained with DAPI (blue).
    • Flow Cytometry - Anti-AHNAK antibody [EM-09] (ab68556)
      Flow Cytometry - Anti-AHNAK antibody [EM-09] (ab68556)
      Overlay histogram showing HeLa cells stained with ab68556 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab68556, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in Hela cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

    References (3)

    Publishing research using ab68556? Please let us know so that we can cite the reference in this datasheet.

    ab68556 has been referenced in 3 publications.

    • Zhang Z  et al. Upregulation of nucleoprotein AHNAK is associated with poor outcome of pancreatic ductal adenocarcinoma prognosis via mediating epithelial-mesenchymal transition. J Cancer 10:3860-3870 (2019). PubMed: 31333803
    • Johnson S  et al. Mass spectrometry analysis reveals differences in the host cell protein species found in pseudotyped lentiviral vectors. Biologicals 52:59-66 (2018). WB . PubMed: 29361371
    • Park JW  et al. AHNAK Loss in Mice Promotes Type II Pneumocyte Hyperplasia and Lung Tumor Development. Mol Cancer Res 16:1287-1298 (2018). PubMed: 29724814

    Customer reviews and Q&As

    Show All Reviews Q&A
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    1-5 of 5 Abreviews or Q&A

    Western blot abreview for Anti-AHNAK antibody [EM-09]

     Poor
    Abreviews
    This product is known to not work in this application or species.
    abreview image
    Application
    Western blot
    Sample
    Rat Cell lysate - whole cell (adipocytes)
    Loading amount
    30 µg
    Specification
    adipocytes
    Gel Running Conditions
    Non-reduced Denaturing (4-12% gradient)
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Feb 01 2011

    Question

    Here are some additional questions from the customer:
    Prepare tissue sections as usual.
    At the usual blocking step, block with serum (from same species as the secondary antibody) for 30 min at room temperature- Does Abcam sell this specific mouse serum?
    Wash 3 X 2 min with PBS Tween 20.
    Incubate sections with an unconjugated AffiniPure Fab fragment Anti-Mouse IgG (H+L) for 1 hr at room temperature, or overnight 4°C . Try this blocking antibody at 0.1 mg/ml although the optimal concentration will need to be assessed by the end user.- Does this IgG come from Abcam?
    Proceed with antibody staining.- If I dont have to order mouse on mouse, which other kit can I order to perform IHC with mouse monoclonal antibody?
    Thank you very much for your continued help.
    Regards,

    Read More
    Answer

    We do sell normal mouse serum, the catalogue number is ab7486 and ab138705.

    The affinity purified antibody we recommend is ab139582; https://www.abcam.com/goat-polyclonal-secondary-antibody-to-mouse-igg-fab-pre-adsorbed-ab139582.html

    We sell compatible IHC kits like ab64264, ab93697 etc.

    https://www.abcam.com/index.html?pageconfig=resource&rid=12906

    Read More

    Question


    I looked at our additional slides this weekend and I think the problem is not with the antibody but with autofluorescence of our cells in the FITC channel that is bleeding through to the RRX channel. With some modifications to the filter settings we should now be able to use the AHNAK antibody with these cells.
    Thanks for your help,

    Read More
    Answer

    Thank you for following up.


    I am happy to hear that the antibody is working as expected. Please do not hesitate to contact me if you have any additional questions. Good luck with your research!

    Read More

    Question

    I tested in this antibody in IF, but saw positive fluorescent signal in the absence of a secondary antibody. Can you confirm that this antibody is not conjugated?

    Read More
    Answer

    Thank you for contacting Abcam regarding ab68556.


    I have followed up with the lab and our shipping/receiving department and we have been unable to determine why this antibody is autofluorescing. I have confirmed that the antibody is not, nor is any version of it conjugated to a fluorophore and other vials of the antibody have been labeled correctly.


    I am wondering how your additional slides looked? Would you like me to send you another vial of this antibody?

    I look forward to your reply so that I may assist you further. Please do not hesitate to contact me if you have any additional questions.

    Read More

    Question

    is there an explanation for the band at 150kDa? could it be a splice variant?

    Read More
    Answer

    Thank you for your patience. The lab sent the following explanation: The 150 kDa band seems to be a fragment arising due to manipulation with the extremely large AHNAK1 protein. Sometimes there are even more fragments observable, as the SDS PAGE / Western blotting application with such huge protein is not easy.    I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information. 

    Read More

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