Human CAT (Catalase) knockout HeLa cell line (ab265250)
Overview
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Product name
Human CAT (Catalase) knockout HeLa cell line
See all Catalase lysates -
Parental Cell Line
HeLa -
Organism
Human -
Mutation description
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 -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
Tested applications
Suitable for: WBmore details -
Biosafety level
2 -
General 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.
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. Based on cell count, seed cells in an appropriate cell culture 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.
4. Incubate the culture at 37°C incubator with 5% CO2. 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.
- A partial media change 24 hours prior to subculture may be helpful to encourage growth, if required.
- Cells should be passaged when they have achieved 80-90% confluence.
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.
We will provide viable cells that proliferate on revival.
Properties
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Number of cells
1 x 106 cells/vial, 1 mL -
Adherent /Suspension
Adherent -
Tissue
Cervix -
Cell type
epithelial -
Disease
Adenocarcinoma -
Gender
Female -
STR Analysis
Amelogenin X D5S818: 11, 12 D13S317: 12, 13.3 D7S820: 8, 12 D16S539: 9, 10 vWA: 16, 18 TH01: 7 TPOX: 8,12 CSF1PO: 9, 10 -
Mycoplasma free
Yes -
Storage instructions
Shipped on Dry Ice. Store in liquid nitrogen. -
Storage buffer
Constituents: 8.7% Dimethylsulfoxide, 2% Cellulose, methyl ether -
Research areas
Target
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Function
Occurs in almost all aerobically respiring organisms and serves to protect cells from the toxic effects of hydrogen peroxide. Promotes growth of cells including T-cells, B-cells, myeloid leukemia cells, melanoma cells, mastocytoma cells and normal and transformed fibroblast cells. -
Involvement in disease
Defects in CAT are the cause of acatalasia (ACATLAS) [MIM:115500]; also known as acatalasemia. This disease is characterized by absence of catalase activity in red cells and is often associated with ulcerating oral lesions. -
Sequence similarities
Belongs to the catalase family. -
Post-translational
modificationsThe N-terminus is blocked. -
Cellular localization
Peroxisome. - Information by UniProt
Associated products
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KO cell lysates
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab265250 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 60 kDa.
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Notes |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 60 kDa. |
Images
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All lanes : Anti-Catalase antibody [EPR20198] - Peroxisome Marker (ab209211) at 1/2000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : CAT knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 60 kDa
Observed band size: 60 kDaLanes 1-2: Merged signal (red and green). Green - ab209211 observed at 60 kDa. Red - loading control ab8245 observed at 37 kDa.
ab209211 Anti-Catalase antibody [EPR20198] 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 ab256859) was used. Wild-type and Catalase knockout samples were subjected to SDS-PAGE. ab209211 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 2000 and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and 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.
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All lanes : Anti-Catalase antibody [EP1929Y] - Peroxisome Marker (ab76024) at 1/10000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : CAT knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 60 kDa
Observed band size: 60 kDaLanes 1-2: Merged signal (red and green). Green - ab76024 observed at 60 kDa. Red - loading control ab8245 observed at 37 kDa.
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 ab256859) was used. Wild-type and Catalase knockout samples were subjected to SDS-PAGE. ab76024 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 10000 and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and 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.
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Allele-1: 4 bp deletion in exon 1.
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Allele-2: 1 bp deletion in exon 1.
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Allele-3: Insertion of the selection cassette in exon 1.
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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.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab265250 has not yet been referenced specifically in any publications.