Human CD63 knockout Hep G2 cell line (ab280796)
Overview
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Product name
Human CD63 knockout Hep G2 cell line -
Parental Cell Line
HepG2 -
Organism
Human -
Passage number
<20 -
Knockout validation
Western Blot (WB) -
Biosafety level
1 -
General notes
Recommended control: Human wild-type HepG2 cell line (ab275467). 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: MEM + 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 and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant 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
Liver -
Cell type
epithelial -
Disease
Hepatocellular Carcinoma -
Gender
Male -
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
This antigen is associated with early stages of melanoma tumor progression. May play a role in growth regulation. -
Tissue specificity
Dysplastic nevi, radial growth phase primary melanomas, hematopoietic cells, tissue macrophages. -
Sequence similarities
Belongs to the tetraspanin (TM4SF) family. -
Cellular localization
Cell membrane. Lysosome membrane. Late endosome membrane. Also found in Weibel-Palade bodies of endothelial cells. Located in platelet dense granules. - Information by UniProt
Images
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All lanes : Anti-CD63 antibody [EPR5702] - Late Endosome Marker (ab134045) at 1/1000 dilution
Lane 1 : Wild-type HepG2 Vehicle Control Brefeldin A (0 u/mL, 24h) cell lysate
Lane 2 : Wild-type HepG2 Treated Brefeldin A (10 u/mL, 24h) cell lysate
Lane 3 : CD63 knockout HepG2 Vehicle Control Brefeldin A (0 u/mL, 24h) cell lysate
Lane 4 : CD63 knockout HepG2 Treated Brefeldin A (10 u/mL, 24h) cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Observed band size: 30 kDa why is the actual band size different from the predicted?Anti-CD63 antibody [EPR5702] (ab134045) staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab134045 was shown to bind specifically to CD63. A band was observed at 30 kDa in wild-type HepG2 cell lysates with no signal observed at this size in CD63 knockout cell line ab280796 (knockout cell lysate ab283828). To generate this image, wild-type and CD63 knockout HepG2 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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 then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab280796 has not yet been referenced specifically in any publications.