Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EP867Y] to Huntingtin - BSA and Azide free
  • Suitable for: WB, IHC-P, Flow Cyt, ICC/IF
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Overview

  • Product name

    Anti-Huntingtin antibody [EP867Y] - BSA and Azide free
    See all Huntingtin primary antibodies

  • Description

    Rabbit monoclonal [EP867Y] to Huntingtin - BSA and Azide free

  • Host species

    Rabbit

  • Tested applications

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

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: SH-SY5Y, HAP1, and HeLa cell lysates. ICC/IF: SKNSH cells. Flow Cyt: SH-SY5Y cells. IHC-P: Human brain tissue.

  • General notes

    ab225573 is the carrier-free version of ab45169. 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

    ab225573 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    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

Images

  • Western blot - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)
    Western blot - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)
    All lanes : Anti-Huntingtin antibody [EP867Y] (ab45169) at 1/10000 dilution

    Lane 1 : Wild-type HeLa cell lysate
    Lane 2 : HTT knockout HeLa cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 348 kDa
    Observed band size: 348 kDa



    This data was developed using the same antibody clone in a different buffer formulation (ab45169).

      Lanes 1- 2: Merged signal (red and green). Green - ab45169 observed at 348 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.

     ab45169 was shown to react with Huntingtin in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab265976 (knockout cell lysate ab256946) was used. Wild-type HeLa and HTT knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab45169 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) overnight at 4°C at a 1 in 10000 dilution and a 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.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)

    Ab45169 staining human Huntingtin in human brain tissue by immunohistochemistry using paraffin embedded tissue.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab45169).

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

  • Immunocytochemistry/ Immunofluorescence - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)
    Immunocytochemistry/ Immunofluorescence - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)

    ICC/IF image of ab45169 stained SKNSH cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab45169, 1/200 dilution) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab45169).

  • Western blot - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)
    Western blot - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)
    All lanes : Anti-Huntingtin antibody [EP867Y] (ab45169) at 1/10000 dilution

    Lane 1 : Wild-type HAP1 whole cell lysate
    Lane 2 : Huntingtin knockout HAP1 whole cell lysate
    Lane 3 : SH-SY-5Y whole cell lysate
    Lane 4 : HeLa whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 348 kDa



    This WB data was generated using the same anti-Huntingtin antibody clone [EP867Y] in a different buffer formulation (cat# ab45169).

    Lanes 1 - 4: Merged signal (red and green). Green - ab45169 observed at 348 kDa. Red - loading control, ab18058, observed at 130 kDa.

    ab45169 was shown to recognize Huntingtin when Huntingtin knockout samples were used, along with additional cross-reactive bands. Wild-type and Huntingtin knockout samples were subjected to SDS-PAGE. ab45169 and ab18058 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C both at 1/10000 dilution. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

  • Flow Cytometry - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)
    Flow Cytometry - Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)

    Overlay histogram showing SH-SY5Y cells stained with ab45169 (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 (ab45169, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) ( 1µg/1x10^6 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in SH-SY5Y cells fixed with 4% paraformaldehyde/permeabilized in 0.1% PBS-Tween used under the same conditions.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab45169).

  • Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)
    Anti-Huntingtin antibody [EP867Y] - BSA and Azide free (ab225573)

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