THOP1 KO cell line available now. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2.
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Key facts
- Cell type
HEK-293T
- Species or organism
Human
- Tissue
Kidney
- Form
Liquid
- Knockout validation
Sanger Sequencing, Western blot
Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2
Recommended products
THOP1 KO cell line available now. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2.
Key facts
- Cell type
HEK-293T
- Species or organism
Human
- Tissue
Kidney
- Form
Liquid
- Knockout validation
Sanger Sequencing, Western blot
- Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2
- Concentration
Properties
- Gene name
THOP1
- Gene editing type
Knockout
- Gene editing method
CRISPR technology
- Knockout validation
Sanger Sequencing, Western blot
Cell culture
- Biosafety level
EU: 2 US: 2
- Viability
~ 80%
- 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 long-term storage conditions
-196°C
Notes
Although we aim to provide customers with a homozygous clone, feasibility will be dependent on the biology of the protein. Should only heterozygous edits be achieved, you will be notified of the outcome and be asked to confirm whether the cell line is acceptable. All clones will be accompanied with DNA sequencing data, and the mutation description.
Recommended control: Human wild-type HEK293T cell line (Human wild-type HEK-293T cell line ab255449). 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.
Cryopreservation cell medium: Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Culture medium: DMEM (High Glucose) + 10% FBS
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 2x10E4 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 2x10E4 cells/cm2 is recommended.
• Cells should be passaged when they have achieved 80-90% confluence.
• Do not allow the cell density to exceed 7x10E4 cells/cm2.
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
- Activity summary
Thimet Oligopeptidase also known as EC 3.4.24.15 or TOP functions as a zinc-dependent metallopeptidase. It breaks down peptides that are up to 20 amino acids long by cleaving peptide bonds. It has a molecular mass of approximately 78 kDa. This enzyme is expressed in various tissues prominently in the brain but also found in the kidneys lungs and testis reflecting its broad physiological roles. TOP primarily exists in the cytosol but can translocate to other cellular compartments under specific conditions.
- Biological function summary
Thimet Oligopeptidase plays a significant role in modulating neuropeptide levels in the nervous system. It does not operate as part of a known complex but instead acts independently in intracellular peptide catabolism. This activity helps regulate processes such as pain modulation and neuroendocrine signaling by modifying peptide-based signaling molecules. Furthermore TOP participates in the degradation of antigenic peptides implicating a role in immune system regulation.
- Pathways
Thimet Oligopeptidase participates in neuropeptide and peptide hormone metabolism where it helps maintain homeostasis by modulating bioactive peptide concentrations. Within these pathways TOP interacts with other proteins such as neprilysin and angiotensin-converting enzyme creating a network that helps regulate cardiovascular and neural functions. This interaction highlights the enzyme's versatile capability to influence multiple signaling cascades.
- Associated diseases and disorders
Thimet Oligopeptidase connects strongly to neurodegenerative diseases such as Alzheimer's and Parkinson's. Research indicates that abnormal activity of TOP contributes to the accumulation of toxic peptide fragments like amyloid-beta in Alzheimer’s disease. This connection brings it into association with proteins like amyloid precursor protein (APP) and tau which are key in such neurodegenerative conditions. The enzyme's regulation of peptidase activity in these contexts highlights its potential as a therapeutic target for disease intervention.
Product promise
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In the unlikely event of one of our products not working as expected, you are covered by our product promise.
Full details and terms and conditions can be found here:
Terms & Conditions.
1 product image
Sanger Sequencing - Human THOP1 (Thimet Oligopeptidase) knockout HEK-293T cell line (ab266421)
Homozygous: 1 bp insertion in exon2
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