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Purified anti-ITCH Antibody

Pricing & Availability
Clone
W19229A (See other available formats)
Regulatory Status
RUO
Other Names
ITCHY, Atrophin-1-interacting protein 4, AIP4, HECT-type E3 ubiquitin transferase Itchy homolog, NFE2-Associated Polypeptide 1, NAPP1, AIF4, ADMFD
Isotype
Rat IgG2b, κ
Ave. Rating
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Product Citations
publications
a. W19229A_Purified_ITCH_Antibody_WB_030323
Whole cell extracts (15 µg total protein) from the indicated cell lines were resolved on a 4-12% Bis-Tris gel, transferred to a PVDF membrane, and probed with 0.25 µg/mL of purified anti-ITCH (clone W19229A) overnight at 4°C. Proteins were visualized by chemiluminescence detection using HRP goat anti-rat IgG (Cat. No. 405405) at a 1:3000 dilution. Direct-Blot™ HRP anti-GAPDH (Cat. No. 607904) was used as a loading control at a 1:50000 dilution. Western-Ready™ ECL substrate premium kit (Cat. No. 426319) was used as a detection agent. Lane M: Molecular weight marker
  • a. W19229A_Purified_ITCH_Antibody_WB_030323
    Whole cell extracts (15 µg total protein) from the indicated cell lines were resolved on a 4-12% Bis-Tris gel, transferred to a PVDF membrane, and probed with 0.25 µg/mL of purified anti-ITCH (clone W19229A) overnight at 4°C. Proteins were visualized by chemiluminescence detection using HRP goat anti-rat IgG (Cat. No. 405405) at a 1:3000 dilution. Direct-Blot™ HRP anti-GAPDH (Cat. No. 607904) was used as a loading control at a 1:50000 dilution. Western-Ready™ ECL substrate premium kit (Cat. No. 426319) was used as a detection agent. Lane M: Molecular weight marker
  • b. W19229A_Purified_ITCH_Antibody_ICC_030323
    Jurkat cells were fixed and permeabilized with ice-cold methanol and blocked with 5% FBS for 1 hour at room temperature. The cells were then stained with 1.25 µg/mL of purified anti-ITCH (clone W19229A), followed by incubation with 2.5 µg/mL of Alexa Fluor® 594 goat anti-rat IgG (Panels A and B) (Cat. No. 405422) for 1 hour at room temperature. Nuclei were counterstained with DAPI (panel B) (Cat. No. 422801), and the image was captured with a 60X objective. Scale bar: 50 µm
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611451 25 µg £83
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611452 100 µg £202
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Description

ITCH, Itchy E3 Ubiquitin Protein Ligase or atrophin-1 interacting protein 4 (AIP4), receives ubiquitin from E2 ubiquitin-conjugating enzymes and transfers them to their target proteins for lysosomal degradation. The degradation regulated by ITCH is involved in a broad array of biological functions, including T helper cell inflammatory signaling, T lymphocyte differentiation, immune response, tumor cell death, and skin keratinocyte differentiation. In animal models, ITCH mutant mice develop spontaneous autoimmune disease symptoms including skin scratching, lymphoproliferation, and chronic pulmonary inflammation. ITCH suppresses IL-4 levels by degrading JUNB and maintains the attenuation of differentiation of T helper cells type 2. To maintain peripheral immune tolerance, ITCH monoubiquitinates protein kinase θ and phospholipase C-γ1 through lysosome degradation near immunological synapse for T cell energy. ITCH modulates trafficking of receptor CXCR4 from the cell surface to the endosome through endocytosis. The transcription factor targets of ITCH, such as JUN, JUNB, TP63, NOTCH1, and GLI1, participate in epidermal keratinocyte development and differentiation. In addition to a role in proteolysis, the polyubiquitination of BRAF by ITCH sustains its activity and elevates the MEK/ERK signaling pathway to promote proliferation and invasion in melanoma cells.

Product Details
Technical Data Sheet (pdf)

Product Details

Verified Reactivity
Human, Mouse
Antibody Type
Monoclonal
Host Species
Rat
Immunogen
Recombinant fragment of human ITCH
Formulation
Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide
Preparation
The antibody was purified by affinity chromatography.
Concentration
0.5 mg/mL
Storage & Handling
The antibody solution should be stored undiluted between 2°C and 8°C.
Application

WB - Quality tested
ICC - Verified
Recommended Usage

Each lot of this antibody is quality control tested by western blotting. For western blotting, the suggested use of this reagent is 0.25 - 1.0 µg/mL. For immunocytochemistry, a concentration range of 1.25 - 5.0 μg/mL is recommended. It is recommended that the reagent be titrated for optimal performance for each application.

Application Notes

For immunocytochemistry (ICC), we suggest fixation with 4% PFA followed by permeabilization with ice-cold methanol or fixation and permeabilization with ice-cold methanol. Fixation with 4% PFA followed by permeabilization with Triton X-100 results in dimmer staining in comparison to the other two methods.

Clone W19229A is not suitable for IHC.

RRID
AB_2936758 (BioLegend Cat. No. 611451)
AB_2936758 (BioLegend Cat. No. 611452)

Antigen Details

Structure
ITCH is a 903 amino-acid protein with a predicted molecular weight of 102.8 kD
Distribution

Ubiquitous; localized to the membrane, cytoplasm, nucleus, trans-Golgi network, endosome, and lysosome

Function
E3 ubiquitin-protein ligase transferring ubiquitin to target protein
Interaction
JUN, JUNB, CXCR4, FYN, ERRB4, protein kinase θ, phospholipase C-γ1 and virus proteins
Cell Type
B cells, Basophils, Dendritic cells, Eosinophils, Lymphocytes, Neutrophils, NK cells, T cells, Th2, Tregs
Biology Area
Adaptive Immunity, Apoptosis/Tumor Suppressors/Cell Death, Cell Biology, Cell Death, Immunology, Innate Immunity, Protein Trafficking and Clearance, Signal Transduction, Stem Cells, Ubiquitin/Protein Degradation
Molecular Family
Endosomal Markers, Enzymes and Regulators, Innate Immune Signaling, Ligases, Lysosomal Markers
Antigen References
  1. Marchese A, et al. 2003. Dev Cell. 5:709-22.
  2. Chastagner P, et al. 2008. PLoS One. 3:e2735
  3. Sundvall M, et al. 2008. Proc Natl Acad Sci U S A. 105:4162-7.
  4. Yin Q, et al. 2019. Nat Commun. 10:1870.

 

Gene ID
83737 View all products for this Gene ID
UniProt
View information about ITCH on UniProt.org
Go To Top Version: 1    Revision Date: 03/03/2023

For Research Use Only. Not for diagnostic or therapeutic use.

 

This product is supplied subject to the terms and conditions, including the limited license, located at www.biolegend.com/terms) ("Terms") and may be used only as provided in the Terms. Without limiting the foregoing, BioLegend products may not be used for any Commercial Purpose as defined in the Terms, resold in any form, used in manufacturing, or reverse engineered, sequenced, or otherwise studied or used to learn its design or composition without express written approval of BioLegend. Regardless of the information given in this document, user is solely responsible for determining any license requirements necessary for user’s intended use and assumes all risk and liability arising from use of the product. BioLegend is not responsible for patent infringement or any other risks or liabilities whatsoever resulting from the use of its products.

 

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This data display is provided for general comparisons between formats.
Your actual data may vary due to variations in samples, target cells, instruments and their settings, staining conditions, and other factors.
If you need assistance with selecting the best format contact our expert technical support team.

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