Validated using a knockout cell line
Recombinant
RabMAb

Anti-Huntingtin antibody [EPR5526] (ab109115)

Overview

  • Product name
    Anti-Huntingtin antibody [EPR5526]
    See all Huntingtin primary antibodies
  • Description
    Rabbit monoclonal [EPR5526] to Huntingtin
  • Tested applications
    Suitable for: Flow Cyt, ICC/IF, WB, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) corresponding to Human Huntingtin aa 1-100.
    Database link: P42858

  • Positive control
    • WB: SH-SY5Y, HeLa, PC-12 and Neuro-2a whole cell lysates and mouse and rat brain lysates. IHC-P: Human cerebral cortex tissue, Human astrocytoma tissue, Mouse testis and Rat testis tissue. ICC/IF: Neuro-2a and SH-SY5Y cell lines. Flow cytometry: SH-SY5Y cell line.
  • General notes

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

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
  • Storage buffer
    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 40% Glycerol, 59% PBS, 0.05% BSA
  • Concentration information loading...
  • Purity
    Protein A purified
  • Clonality
    Monoclonal
  • Clone number
    EPR5526
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab109115 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 1/250.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

ICC/IF 1/1000.
WB 1/5000. Detects a band of approximately 348 kDa (predicted molecular weight: 348 kDa).
IHC-P 1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Target

  • Function
    May play a role in microtubule-mediated transport or vesicle function.
  • Tissue specificity
    Expressed in the brain cortex (at protein level). Widely expressed with the highest level of expression in the brain (nerve fibers, varicosities, and nerve endings). In the brain, the regions where it can be mainly found are the cerebellar cortex, the neocortex, the striatum, and the hippocampal formation.
  • Involvement in disease
    Defects in HTT are the cause of Huntington disease (HD) [MIM:143100]. HD is an autosomal dominant neurodegenerative disorder characterized by involuntary movements (chorea), general motor impairment, psychiatric disorders and dementia. Onset of the disease occurs usually in the third or fourth decade of life and symptoms progressively worsen leading to death in 10 to 20 years. Onset and clinical course depend on the degree of poly-Gln repeat expansion, longer expansions resulting in earlier onset and more severe clinical manifestations. HD affects 1 in 10,000 individuals of European origin. Neuropathology of Huntington disease displays a distinctive pattern with loss of neurons, especially in the caudate and putamen (striatum).
  • Sequence similarities
    Belongs to the huntingtin family.
    Contains 10 HEAT repeats.
  • Domain
    The N-terminal Gln-rich and Pro-rich domain has great conformational flexibility and is likely to exist in a fluctuating equilibrium of alpha-helical, random coil, and extended conformations.
  • Post-translational
    modifications
    Cleaved by apopain downstream of the polyglutamine stretch. The resulting N-terminal fragment is cytotoxic and provokes apoptosis.
    Forms with expanded polyglutamine expansion are specifically ubiquitinated by SYVN1, which promotes their proteasomal degradation.
  • Cellular localization
    Cytoplasm. Nucleus. The mutant Huntingtin protein colocalizes with AKAP8L in the nuclear matrix of Huntington's disease neurons.
  • Information by UniProt
  • Database links
  • Alternative names
    • AI256365 antibody
    • C430023I11Rik antibody
    • HD antibody
    • HD protein antibody
    • HD_HUMAN antibody
    • HDH antibody
    • HTT antibody
    • Huntingtin antibody
    • HUNTINGTON CHOREA antibody
    • Huntington disease protein antibody
    • Huntington's disease protein homolog antibody
    • IT 15 antibody
    • IT15 antibody
    • OTTMUSP00000026909 antibody
    • ZHD antibody
    see all

Images



  • Predicted band size : 348 kDa

    Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
    Lane 2: HTT knockout HAP1 whole cell lysate (20 µg)
    Lane 3: SH-SY5Y whole cell lysate (20 µg)
    Lane 4: HeLa whole cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab109115 observed at 348 kDa. Red - loading control, ab18058, observed at 130 kDa.

    ab109115 was shown to specifically recognize HTT in wild-type HAP1 cells along with additional cross-reactive bands. No band was observed when HTT knockout samples were exmined. Wild-type and HTT knockout samples were subjected to SDS-PAGE. Unpurified ab109115 and ab18058 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/10,000 dilution and 1/10,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

  • All lanes : Anti-Huntingtin antibody [EPR5526] (ab109115) at 1/5000 dilution (purified)

    Lane 1 : SH-SY5Y (Human neuroblastoma from bone marrow cells) whole cell lysate
    Lane 2 : HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate
    Lane 3 : PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate
    Lane 4 : Neuro-2a (Mouse neuroblastoma cells) whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution

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


    Exposure time : 1 second

    Blocking and Diluting buffer and concentration:  5% NFDM /TBST

  • Immunohistochemical analysis of paraffin-embedded Human cerebral cortex tissue labeling Huntingtin with purified ab109115 at 1/100 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500. Counter stained with Hematoxylin. Nuclear staining on neuron of human cerebral cortex was observed.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized

    Neuro-2a (Mouse neuroblastoma cells) cells labeling Huntingtin with purified ab109115 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

    Confocal image showing nuclear and cytoplasmic staining on Neuro-2a cell line.

    The nuclear counter stain is DAPI (blue).

    Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

    The negative controls are as follows:
    1. ab191472 at 1/1000 dilution followed by ab150120 (Alexa Fluor® 594 Goat anti-Mouse secondary) at 1/1000 dilution.
    2. ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor® 488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

  • All lanes : Anti-Huntingtin antibody [EPR5526] (ab109115) at 1/50000 dilution (purified)

    Lane 1 : Mouse brain lysate
    Lane 2 : Rat brain lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution

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


    Exposure time : 30 seconds

    Blocking and Diluting buffer and concentration: 5% NFDM /TBST

  • Immunohistochemical analysis of paraffin-embedded Human astrocytoma labeling Huntingtin with purified ab109115 at 1/100 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500. Counter stained with Hematoxylin. Nuclear staining on cancer cells of astrocytoma.

  • Flow cytometric analysis of 4% paraformaldehyde-fixed SH-SY5Y (Human neuroblastoma from bone marrow cells) cells labeling Huntingtin with purified ab109115 at 1/250 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (FITC) at 1/500 dilution was used as the secondary antibody.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized

    SH-SY5Y (Human neuroblastoma from bone marrow cells) cells labeling Huntingtin with purified ab109115 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

    Confocal image showing nuclear and cytoplasmic staining on SH-SY5Y cell line.

    The nuclear counter stain is DAPI (blue).

    Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (Alexa Fluor® 594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

    The negative controls are as follows:
    1. ab191472 at 1/1000 dilution followed by ab150120 (Alexa Fluor® 594 Goat anti-Mouse secondary) at 1/1000 dilution.
    2. ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor® 488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

  • Immunohistochemical analysis of paraffin-embedded Mouse testis labeling Huntingtin with purified ab109115 at 1/100 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500. Counter stained with Hematoxylin. Cytoplasmic staining on spermatogenic cells of mouse testis.

  • Immunohistochemical analysis of paraffin-embedded Rat testis labeling Huntingtin with purified ab109115 at 1/100 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500. Counter stained with Hematoxylin. Weak cytoplasmic staining on spermatogenic cells of rat testis.

References

This product has been referenced in:
  • Nakasuji T  et al. Complementary Critical Functions of Zfy1 and Zfy2 in Mouse Spermatogenesis and Reproduction. PLoS Genet 13:e1006578 (2017). WB . Read more (PubMed: 28114340) »
  • Schut MH  et al. Effect of post-mortem delay on N-terminal huntingtin protein fragments in human control and Huntington disease brain lysates. PLoS One 12:e0178556 (2017). Read more (PubMed: 28570578) »

See all 5 Publications for this product

Customer reviews and Q&As

Application
Immunoprecipitation
Sample
Mouse Tissue lysate - whole (Brain)
Total protein in input
1400 µg
Immuno-precipitation step
Other - Dynabeads
Specification
Brain
Username

Svitlana Yablonska

Verified customer

Submitted Apr 19 2017

Application
Western blot
Sample
Escherichia coli Purified protein (Purified protein)
Gel Running Conditions
Reduced Denaturing (10% gel)
Loading amount
2 µg
Treatment
Protease cleavage 3hrs
Specification
Purified protein
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Ms. Ya Zhou

Verified customer

Submitted Jun 28 2016

Application
Western blot
Sample
Bacteria Cell lysate - whole cell (BL21)
Gel Running Conditions
Non-reduced Denaturing (10% gel)
Loading amount
10 µg
Treatment
IPTG 1mM 16h
Specification
BL21
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted Jun 27 2016

Thank you for contacting us.

The immunogen is from N-term from amino acid XXXX of P42858.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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