Validated using a knockout cell line

Anti-Islet 1 antibody [1H9] (ab86472)

Reviews (1)Q&A (3)References (8)

Overview

  • Product name

    Anti-Islet 1 antibody [1H9]
    See all Islet 1 primary antibodies

  • Description

    Mouse monoclonal [1H9] to Islet 1

  • Host species

    Mouse

  • Specificity

    In western blot, we observe a specific band at ~40kDa which is not seen in KO cell lines. A cross-reactive band was observed in the wild-type and knockout cells. The additional band below this band of interest is seen at ~38kDa in both the WT and KO cells. We are unsure as to the identity of these extra bands.

  • Tested applications

    Suitable for: Flow Cyt, WB, ELISA, IHC-P, ICC/IFmore details

  • Species reactivity

    Reacts with: Human

  • Immunogen

    Recombinant fragment corresponding to Human Islet 1 aa 150-349. Purified recombinant fragment of Human Islet 1 expressed in E. Coli
    Database link: P61371

  • Positive control

    • Transfected HEK293 cell lysate. Human lung cancer tissue and cervical carcinoma tissue.

  • General notes

    This product was changed from ascites to supernatant. Lot no’s high than GR141006-29 are from Tissue Culture Supernatant

Properties

Applications

Our Abpromise guarantee covers the use of ab86472 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
Flow Cyt 1/50 - 1/100.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
WB 1/500 - 1/2000. Predicted molecular weight: 39 kDa.
ELISA 1/10000.
IHC-P 1/200 - 1/1000.
ICC/IF 1/200 - 1/1000.

Target

  • Function

    Binds to one of the cis-acting domain of the insulin gene enhancer.

  • Tissue specificity

    Expressed in subsets of neurons of the adrenal medulla and dorsal root ganglion, inner nuclear and ganglion cell layers in the retina, the pineal and some regions of the brain.

  • Sequence similarities

    Contains 1 homeobox DNA-binding domain.
    Contains 2 LIM zinc-binding domains.

  • Cellular localization

    Nucleus.
  • Information by UniProt

  • Database links

    • Alternative names

      • Insulin gene enhancer protein ISL 1 antibody
      • Insulin gene enhancer protein ISL-1 antibody
      • Insulin related protein antibody
      • ISL 1 antibody
      • ISL LIM homeobox 1 antibody
      • ISL1 antibody
      • ISL1 transcription factor LIM homeodomain antibody
      • ISL1 transcription factor, LIM/homeodomain (islet 1) antibody
      • ISL1 transcription factor, LIM/homeodomain antibody
      • ISL1_HUMAN antibody
      • Islet-1 antibody
      • Islet1 antibody
      see all

    Images

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Islet 1 antibody [1H9] (ab86472)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Islet 1 antibody [1H9] (ab86472)
      ab86472 at a 1/200 dilution staining Islet 1 in human lung cancer tissue. DAB staining shows nuclear localisation.
    • Western blot - Anti-Islet 1 antibody [1H9] (ab86472)
      Western blot - Anti-Islet 1 antibody [1H9] (ab86472)
      All lanes : Anti-Islet 1 antibody [1H9] (ab86472) at 1/500 dilution

      Lane 1 : Wild-type HAP1 whole cell lysate
      Lane 2 : ISL1 (Islet 1) knockout HAP1 whole cell lysate
      Lane 3 : SH-SY5Y whole cell lysate
      Lane 4 : HepG2 whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Predicted band size: 39 kDa
      Observed band size: 40 kDa
      why is the actual band size different from the predicted?



      Lanes 1 - 4: Merged signal (red and green). Green - ab86472 observed at 40 kDa. Red - loading control, ab176560, observed at 50 kDa.

      ab86472 was shown to recognize Islet 1 in wild-type HAP1 cells as signal was lost at the expected MW in Islet 1 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and Islet 1 knockout samples were subjected to SDS-PAGE. ab86472 and ab176560 (Rabbit anti-alpha Tubulin loading control) were incubated overnight at 4°C at 1/500 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

    • Western blot - Anti-Islet 1 antibody [1H9] (ab86472)
      Western blot - Anti-Islet 1 antibody [1H9] (ab86472)
      Anti-Islet 1 antibody [1H9] (ab86472) at 1/500 dilution + HEK293 cells transfected with full length Islet 1 (aa1-349)-hIgGFc

      Predicted band size: 39 kDa



      Note: The molecular weight is larger than expected, as the recombinant protein is not the endogenouse protein, but rather the fc-fusion protein

    • Immunocytochemistry/ Immunofluorescence - Anti-Islet 1 antibody [1H9] (ab86472)
      Immunocytochemistry/ Immunofluorescence - Anti-Islet 1 antibody [1H9] (ab86472)

      Confocal immunofluorescence analysis of HEK293 cells trasfected with full-length Islet 1-hIgGFc using ab86472 at 1/150 dilution (green). Blue: DRAQ5 fluorescent DNA dye.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Islet 1 antibody [1H9] (ab86472)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Islet 1 antibody [1H9] (ab86472)
      ab86472 at a 1/200 dilution staining Islet 1 in human cervical carcinoma tissue. DAB staining shows nuclear localization.
    • Flow Cytometry - Anti-Islet 1 antibody [1H9] (ab86472)
      Flow Cytometry - Anti-Islet 1 antibody [1H9] (ab86472)

      Overlay histogram showing SH-SY5Y cells stained with ab86472 (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 (ab86472, 1/50 dilution) 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/1x10^6 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive result in SH-SY5Y cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween for 20 min used under the same conditions.

    References

    This product has been referenced in:

    • Hernandez I  et al. Short-term hypoxia improves early cardiac progenitor cell function in vitro. Am J Stem Cells 7:1-17 (2018). Flow Cyt ; Human . Read more (PubMed: 29531855) »
    • Konukiewitz B  et al. Pancreatic neuroendocrine carcinomas reveal a closer relationship to ductal adenocarcinomas than to neuroendocrine tumors G3. Hum Pathol 77:70-79 (2018). IHC-P ; Human . Read more (PubMed: 29596894) »
    See all 8 Publications for this product

    Publishing research using ab86472? Please let us know so that we can cite the reference in this datasheet.

    Customer reviews and Q&As

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    1-4 of 4 Abreviews or Q&A

    Immunohistochemistry (Frozen sections) abreview for Anti-Islet 1 antibody [1H9]

     Good
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Frozen sections)
    Blocking step
    Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1.5% · Temperature: 37°C
    Sample
    Dog Tissue sections (Heart)
    Specification
    Heart
    Permeabilization
    Yes - 0.5% Tween80 PBS wash
    Fixative
    Paraformaldehyde
    Read More

    Abcam user community

    Verified customer

    Submitted Oct 13 2014

    Question

    Our protocol for Islet 1 staining: 1/800 titer, Ventana Benchmark XT staining platform - CC1 32 minutes, primary antibody 32 minutes @37C, Ventana UltraView DAB detection kit I think credit would be best as we just ordered/received 4 vials of 1 lot that appears to work.

    Read More
    Answer

    Thank you for contacting us. I am sorry that this antibody did not perform as stated on the datasheet. I have asked our accounting department to issue a credit note for you, which can be redeemed against the invoice of a future order by passing it on to your purchasing department. To avoid confusion, please ensure your accounts department is aware of how the credit note is being used. If you have questions on how to use the credit note, please contact our accounting department. Our accounting department can be contacted by email at us.credits@abcam.com or by telephone using the information at the Contact Us link in the top right corner of our website. Please refer to the credit note ID in any correspondence with our accounting department. I hope this experience will not prevent you from purchasing other products from us in the future. Our Scientific Support team is always at your service, should you require further expert advice.

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    Question

     Hello - Hopefully you can help us with some Islet 1 antibody lot issues, we have been working up the Islet 1 in a development setting on paraffin tissue for IHC testing.  We ordered some in August of 2010 and it produced acceptable results.  Then we reordered some in July of 2011 received two lots, both of which produced a lot of nonspecific staining.  Not knowing if it was a protocol issue or not we reordered again a week ago and received another lot that has staining similar to the first lot that we received.  We believe there was some sort of an issue with these lots - are you aware of any problems with these lots?

    Read More
    Answer

    Thank you for contacting Abcam regarding ab86472. I am sorry that you are experiencing difficulties with some lots of this antibody in IHC. I have checked our current stock and currently we have a new lot available. I would be happy to send this as a replacement for the 4 vials that did not work from previous lots. Alternatively, I am happy to provide a credit. Would you also please provide a summary of the protocol you tested with this antibody? This will allow us to investigate this problem further. Thank you in advance for your cooperation so that we may continue to provide high quality antibodies. 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 or concerns.

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    Question

    Wir haben von ihnen einen Antikörper gegen Islet1 geliefert bekommen. Ein früherer Antikörper von Islet1 war bei uns ohne Probleme gelaufen. Dieser Antikörper - ab86472 - Lot.No. GR14734 geht aber leider nicht. Wir haben diesen Antikörper auch bei einen Referenzlabor (gebrauchen auch den geichen Antikörper von Abcam) testen lassen -ohne Erfolg.   Ich bitte sie uns schnellstmöglich einen Ersatz für diese fehlerhafte Lot.No zu schicken.

    Read More
    Answer

    Vielen Dank fuer Ihre email. Es tut mir sehr leid zu hoeren, dass das aktuelle Lot fuer ab86472 Ihnen Probleme bereitet. Eine Ersatzlieferung waere auf jeden Fall moeglich, falls der Antikoerper in getesteten Anwendungen (ELISA, ICC/IF, IHC-P, WB) und Spezien (Hu) verwendet wurde. Koennten Sie mir dazu bitte die folgenden experimentellen Details mitteilen: 1) In welcher Anwendung/Technik wurder der AK verwendet? 2) Was genau ist das Problem? 3) Welche Proben (Spezie, Probenart) wurden benutzt und wie wurden diese hergestellt oder vorbereitet? 4) Welche AK verduennung und Inkubationszeit wurde angewendet? 5) Welche Art von Fixierung, Blockierung und Detektion wurde benutzt? Diese Informationen werden uns mit der Qualitaetskontrolle und Begutachtung unserer Produkte weiterhelfen. Bitte schicken Sie mir diese Informationen so schnell wie moeglich, damit wir das Problem zuegig fuer Sie loesen koennen. Haben Sie vielen Dank im voraus! Ich freue mich wieder von Ihnen zu hoeren!

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