CAT KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 1 and 4 bp deletion in exon 1 and Insertion of the selection cassette in exon 1.
CAS1, CATA_HUMAN, CS 1, Catalase, MGC138422, MGC138424
CAT KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 1 and 4 bp deletion in exon 1 and Insertion of the selection cassette in exon 1.
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.
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.
Catalase also known as CAT is an enzyme that catalyzes the decomposition of hydrogen peroxide into water and oxygen. This enzyme has a molecular weight of approximately 240 kDa and typically forms a tetramer. Catalase mainly resides in peroxisomes functioning as a peroxisome marker. It expresses abundantly in the liver kidneys and erythrocytes where it plays significant roles in cellular protection against oxidative damage. The presence of Catalase makes it an ideal candidate for use in peroxisome staining and peroxisome image analysis in research.
Catalase contributes to antioxidant defense by breaking down hydrogen peroxide preventing cellular damage. In peroxisomes it works alongside other peroxisomal enzymes to maintain cell health and metabolic regulation. Catalase does not form a complex but interacts closely with other enzymes like superoxide dismutase which dismutates superoxide radicals into less harmful substances. Increased Catalase activity levels can be measured using Catalase activity kits and activity assays allowing us to learn about peroxisome function.
Hydrogen peroxide removal by Catalase is vital in the reactive oxygen species (ROS) metabolic process and plays a part in the cellular response to oxidative stress. Catalase interacts with the glutathione peroxidase pathway safeguarding cells from oxidative stress-related damage. Superoxide dismutase works synergistically with Catalase transforming superoxide anions into hydrogen peroxide before its decomposition by Catalase. These activities highlight the essential role Catalase plays in protecting cells from oxidative stress damage.
Catalase relates to conditions like acatalasemia and diabetes. Acatalasemia a condition caused by a deficiency of Catalase increases the risk of developing diabetes and other oxidative stress-related diseases. Mutations in the CAT gene can lead to decreased Catalase activity contributing to the onset of these conditions. Additionally Catalase works with other proteins like glutathione peroxidase in mitigating the effects of oxidative stress with deficiencies potentially exacerbating complications in diabetes management.
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Anti-Catalase antibody [EP1929Y] - Peroxisome Marker ab76024 Anti-Catalase antibody [EP1929Y] - Peroxisome Marker was shown to specifically react with Catalase in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265250 (knockout cell lysate Human CAT (Catalase) knockout HeLa cell lysate ab256859) was used. Wild-type and Catalase knockout samples were subjected to SDS-PAGE. Anti-Catalase antibody [EP1929Y] - Peroxisome Marker ab76024 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4° at 1 in 10000 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 20000 dilution for 1 hour at room temperature before imaging.
All lanes: Western blot - Anti-Catalase antibody [EP1929Y] - Peroxisome Marker (Anti-Catalase antibody [EP1929Y] - Peroxisome Marker ab76024) at 1/10000 dilution
Lane 1: Wild-type HeLa cell lysate at 20 µg
Lane 2: CAT knockout HeLa cell lysate at 20 µg
Lane 2: Western blot - Human CAT (Catalase) knockout HeLa cell line (ab265250)
Performed under reducing conditions.
Predicted band size: 60 kDa
Observed band size: 60 kDa
Anti-Catalase antibody [EPR20198] - Peroxisome Marker ab209211 Anti-Catalase antibody [EPR20198] - Peroxisome Marker was shown to specifically react with Catalase in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265250 (knockout cell lysate Human CAT (Catalase) knockout HeLa cell lysate ab256859) was used. Wild-type and Catalase knockout samples were subjected to SDS-PAGE. Anti-Catalase antibody [EPR20198] - Peroxisome Marker ab209211 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4° at 1 in 2000 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 20000 dilution for 1 hour at room temperature before imaging.
All lanes: Western blot - Anti-Catalase antibody [EPR20198] - Peroxisome Marker (Anti-Catalase antibody [EPR20198] - Peroxisome Marker ab209211) at 1/2000 dilution
Lane 1: Wild-type HeLa cell lysate at 20 µg
Lane 2: CAT knockout HeLa cell lysate at 20 µg
Lane 2: Western blot - Human CAT (Catalase) knockout HeLa cell line (ab265250)
Performed under reducing conditions.
Predicted band size: 60 kDa
Observed band size: 60 kDa
Representative images of CAT 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.
Allele-1: 4 bp deletion in exon 1.
Allele-2: 1 bp deletion in exon 1.
Allele-3: Insertion of the selection cassette in exon 1.
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