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ITCH KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 7.

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Images

Western blot - Human ITCH (AIP4) knockout HeLa cell line (AB265338), expandable thumbnail
  • Sanger Sequencing - Human ITCH (AIP4) knockout HeLa cell line (AB265338), expandable thumbnail

Key facts

Cell type
HeLa
Species or organism
Human
Tissue
Cervix
Form
Liquid
Knockout validation
Sanger Sequencing, Western blot
Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 7

Alternative names

ADMFD, AIF4, Aip-4, Atrophin-1-interacting protein 4, E3 ubiquitin-protein ligase Itchy homolog, EC 6.3.2, ITCH_HUMAN, Itchy E3 ubiquitin protein ligase, Itchy E3 ubiquitin protein ligase homolog, Itchy E3 ubiquitin protein ligase homolog mouse, Itchy E3 ubiquitin protein ligase, mouse, homolog of, Itchy homolog E3 ubiquitin protein ligase, Itchy mouse homolog E3 ubiquitin protein ligase, NAPP1, NFE2-associated polypeptide 1, Ubiquitin protein ligase ITCH, dJ468O1.1, dJ468O1.1 (atrophin 1 interacting protein 4 (AIP4)), dJ468O1.1 atrophin 1 interacting protein 4 AIP4

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ITCH KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 7.

Key facts

Cell type
HeLa
Form
Liquid
Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 7
Disease
Adenocarcinoma
Concentration
Loading...

Properties

Gene name
ITCH
Gene editing type
Knockout
Gene editing method
CRISPR technology
Knockout validation
Sanger Sequencing, Western blot
Zygosity
Homozygous

Quality control

STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX

Cell culture

Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female

Handling procedures

Initial handling guidelines

Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.

1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method seed all remaining cells into a T25.
4. Incubate the culture at 37°C incubator with 5% CO2. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.
5. Once confluent passage into an appropriate flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.

Subculture guidelines
  • All seeding densities should be based on cell counts gained by established methods.
  • A guide seeding density of 2x104 cells/cm2 is recommended.
  • Cells should be passaged when they have achieved 80-90% confluence.
Culture medium
DMEM (High Glucose) + 10% FBS
Cryopreservation medium
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.

Storage

Shipped at conditions
Dry Ice
Appropriate short-term storage conditions
-196°C
Appropriate long-term storage conditions
-196°C

Notes

Recommended control: Human wild-type HeLa cell line (ab255928). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.

We will provide viable cells that proliferate on revival.

This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.
Activity summary

ITCH also known as AIP4 is an E3 ubiquitin ligase with a molecular mass of approximately 108 kDa. The protein primarily functions in the ubiquitination process which regulates protein degradation via the proteasome. ITCH is known for its role in the post-translational modification of proteins through ubiquitin tagging facilitating the promotion or suppression of signal transduction pathways. Expression of ITCH occurs in various tissues including lymphocytes and epithelial cells indicating its diverse functional roles in different physiological contexts.

Biological function summary

ITCH/AIP4 is integral in cellular regulatory mechanisms. It forms part of a sophisticated complex involved in immune response modulation cell cycle regulation and apoptosis. ITCH influences T-cell receptor signaling by controlling the degradation of proteins involved in these critical pathways. Through the ubiquitination cascade ITCH maintains cellular homeostasis participating in the intricate balance of protein synthesis and degradation necessary for normal cellular function.

Pathways

ITCH/AIP4 plays essential roles in the T-cell receptor and Wnt signaling pathways. It interacts with other proteins such as NEDD4 and CBL which are also E3 ubiquitin ligases to modulate signaling cascades involved in immune responses and cellular growth. In the Wnt pathway ITCH regulates the stability of Dishevelled proteins thereby affecting cellular proliferation and differentiation processes. This involvement in multiple pathways highlights its regulatory versatility and importance in maintaining cellular balance.

Associated diseases and disorders

ITCH/AIP4 is associated with autoimmune diseases and cancer. Its dysfunction can lead to abnormal immune responses contributing to conditions such as systemic lupus erythematosus. Moreover the protein's role in oncogenesis is linked to its ability to modify tumor suppressor and oncoprotein levels including p53 and c-Jun. Disruption in ITCH's function can result in unchecked cell growth and survival emphasizing its potential as a therapeutic target in cancer treatment.

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Product protocols

For this product, it's our understanding that no specific protocols are required. You can:

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