CBSL KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 11 bp deletion in exon 5 and 1 bp insertion in exon 5.
Images
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, 11 bp deletion in exon 5 and 1 bp insertion in exon 5
Alternative names
AI047524, AI303044, Beta-thionase, CBS_HUMAN, Cbs cystathionine beta-synthase, Cystathionine beta-synthase, EC 4.2.1.22, HIP 4, MGC18856, MGC18895, MGC37300, Methylcysteine synthase, OTTHUMP00000109416, OTTHUMP00000109418, Serine sulfhydrase
Recommended products
CBSL KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 11 bp deletion in exon 5 and 1 bp insertion in exon 5.
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, 11 bp deletion in exon 5 and 1 bp insertion in exon 5
- Antibiotic resistance
- Puromycin 1µg/mL
- Disease
- Adenocarcinoma
- Concentration
Properties
- Gene name
- CBSL
- Gene editing type
- Knockout
- Gene editing method
- CRISPR technology
- Knockout validation
- Sanger Sequencing, Western blot
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
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
Cystathionine beta-synthase (CBS) is an enzyme that catalyzes the conversion of homocysteine and serine into cystathionine. It is sometimes referred to as CBS monelyne or cystathionine synthase. CBS is a heme-containing protein with a molecular mass of approximately 63 kDa. It is expressed widely in tissues including the liver brain and kidney. The activity of CBS is regulated by various factors including the availability of cofactors like pyridoxal 5'-phosphate and adenosine triphosphate (ATP).
- Biological function summary
The enzyme CBS plays a role in the transsulfuration pathway where it helps in the metabolism of homocysteine. CBS is a part of a larger complex in some tissues where it interacts with other enzymes involved in sulfur amino acid metabolism. This function is important in maintaining cellular sulfur amino acid balance and protecting cells from oxidative stress caused by elevated levels of homocysteine.
- Pathways
CBS functions in the transsulfuration pathway which connects the methionine cycle and the synthesis of glutathione. This pathway is essential for detoxification and antioxidant protection. CBS interrelates with other proteins like cystathionine gamma-lyase (CGL) within this pathway. The interplay between CBS and CGL ensures the conversion of homocysteine to cysteine which is a precursor for the synthesis of glutathione.
- Associated diseases and disorders
CBS deficiency is linked to homocystinuria a disorder characterized by high levels of homocysteine in the blood and urine. Symptoms include cardiovascular problems and developmental delays. Furthermore alterations in CBS activity relate to cardiovascular diseases due to hyperhomocysteinemia. Within these conditions CBS interacts with other proteins involved in homocysteine metabolism such as methionine synthase which also contributes to the complexity of these disorders.
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6 product images
Western blot - Human CBS knockout HeLa cell line (ab264950)
Lanes 1-3: Merged signal (red and green). Green - Anti-CBS antibody [EPR8579] ab140600 observed at 70 kDa. Red - loading control, Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 37 kDa.
Anti-CBS antibody [EPR8579] ab140600 Anti-CBS antibody [EPR8579] was shown to specifically react with CBS in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264950 (knockout cell lysate Human CBS knockout HeLa cell lysate ab257203) was used. Wild-type and CBS knockout samples were subjected to SDS-PAGE. Anti-CBS antibody [EPR8579] ab140600 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.
All lanes: Western blot - Anti-CBS antibody [EPR8579] (Anti-CBS antibody [EPR8579] ab140600) at 1/1000 dilution
Lane 1: Wild-type HeLa cell lysate at 20 µg
Lane 2: CBS knockout HeLa cell lysate at 20 µg
Lane 2: Western blot - Human CBS knockout HeLa cell line (ab264950)
Lane 3: Raji cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 61 kDa
Observed band size: 70 kDa
Western blot - Human CBS knockout HeLa cell line (ab264950)
Lanes 1-3: Merged signal (red and green). Green - Anti-CBS antibody [EPR8578] ab131155 observed at 70 kDa. Red - loading control, Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 37 kDa.
Anti-CBS antibody [EPR8578] ab131155 Anti-CBS antibody [EPR8578] was shown to specifically react with CBS in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264950 (knockout cell lysate Human CBS knockout HeLa cell lysate ab257203) was used. Wild-type and CBS knockout samples were subjected to SDS-PAGE. Anti-CBS antibody [EPR8578] ab131155 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.
All lanes: Western blot - Anti-CBS antibody [EPR8578] (Anti-CBS antibody [EPR8578] ab131155) at 1/1000 dilution
Lane 1: Wild-type HeLa cell lysate at 20 µg
Lane 2: CBS knockout HeLa cell lysate at 20 µg
Lane 2: Western blot - Human CBS knockout HeLa cell line (ab264950)
Lane 3: Raji cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 61 kDa
Observed band size: 70 kDa
Cell Culture - Human CBS knockout HeLa cell line (ab264950)
Representative images of CBS knockout HeLa cells, low and high confluency examples (top left and right respectively) and wild-type HeLa cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using a EVOS XL Core microscope.
Sandwich ELISA - Human CBS knockout HeLa cell line (ab264950)
Sandwich ELISA of Human CBS Antibody Pair - BSA and Azide free ab312869 with the capture antibody dilution at 2 µg/mL and detector antibody dilution at 0.5 µg/mL.
Interpolated concentrations of native CBS in human control wild type HeLa cell and CBS knockout HeLa cell based on 25 µg/mL extract loads. The concentrations of CBS were measured in duplicate and interpolated from the CBS standard curve and corrected for sample dilution. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean CBS concentration was determined to be 4321.4 pg/mL in wild type HeLa extract (Human wild-type HeLa cell line ab255448) and undetectable in CBS knockout HeLa extract (Human CBS knockout HeLa cell line ab264950).
Sanger Sequencing - Human CBS knockout HeLa cell line (ab264950)
Allele-2: 1 bp insertion in exon 5.
Sanger Sequencing - Human CBS knockout HeLa cell line (ab264950)
Allele-1: 11 bp deletion in exon 5.
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