Human FGF21 knockout HeLa cell line (ab265974)
Overview
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Product name
Human FGF21 knockout HeLa cell line -
Parental Cell Line
HeLa -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2 -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
Tested applications
Suitable for: WB, Sanger Sequencingmore 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.
Properties
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Number of cells
1 x 106 cells/vial, 1 mL -
Viability
~80% -
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 -
Antibiotic resistance
Puromycin 1.00µg/ml -
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
Stimulates glucose uptake in differentiated adipocytes via the induction of glucose transporter SLC2A1/GLUT1 expression (but not SLC2A4/GLUT4 expression). Activity requires the presence of KLB. -
Sequence similarities
Belongs to the heparin-binding growth factors family. -
Cellular localization
Secreted. - 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 ab265974 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: 22 kDa.
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Sanger Sequencing |
Use at an assay dependent concentration.
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Notes |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 22 kDa. |
Sanger Sequencing
Use at an assay dependent concentration. |
Images
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All lanes : Anti-FGF21 antibody [EPR8314(2)] (ab171941) at 1/1000 dilution
Lane 1 : Wild-type HeLa Treated BFA (5 ug/mL, 6 h) cell lysate at 20 µg
Lane 2 : FGF21 knockout HeLa Treated BFA (5 ug/mL, 6 h) cell lysate at 20 µg
Lane 3 : Wild-type HeLa Vehicle Control BFA (0 ug/mL, 6 h) cell lysate at 20 µg
Lane 4 : FGF21 knockout HeLa Vehicle Control BFA (0 ug/mL, 6 h) cell lysate at 20 µg
Lane 5 : HepG2 Treated BFA (5 ug/mL, 6 h) cell lysate at 20 µg
Lane 6 : HepG2 cell lysate at 20 µg
Lane 7 : Empty cell lysate at 0 µg
Lane 8 : Human Liver cell lysate at 10 µg
Lane 9 : Mouse Spleen cell lysate at 10 µg
Performed under reducing conditions.
Predicted band size: 22 kDa
Observed band size: 21 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-FGF21 antibody [EPR8314(2)] staining at 1/1000 dilution, shown in black; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab171941 was shown to bind specifically to FGF21. A band was observed at 21 kDa in treated wild-type HeLa cell lysates with no signal observed at this size in FGF21 knockout cell line ab265974 (knockout cell lysate ab256915). To generate this image, wild-type and FGF21 knockout HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times before development with Optiblot (ECL reagent ab133456) and imaged with 16 minutes exposure time. Secondary antibodies used were HRP conjugated Goat anti-Rabbit (H+L) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
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Homozygous: 1 bp insertion in exon 2.
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Representative images of FGF21 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)
ab265974 has not yet been referenced specifically in any publications.