Human CD47 knockout HEK-293T cell line
- Advanced Validation
- What is this?
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(1 Publication)
- ICC
Lab
Immunocytochemistry - Human CD47 knockout HEK-293T cell line (AB266324)
ab213079 staining CD47 in wild-type HEK293 cells (top panel) and CD47 knockout HEK293 cells (bottom panel) (ab266324). The cells were fixed with 100% methanol (5 min) then permeabilized with 0.1% Tween for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab213079 at 0.4 μg/ml concentration and ab6046 (Rabbit polyclonal to beta Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to mouse IgG (Alexa Fluor® 488) (ab150117) at 2 μg/ml (shown in green) and a goat secondary antibody to rabbit IgG (Alexa Fluor® 594) (ab150080) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - Human CD47 knockout HEK-293T cell line (AB266324)
Immunocytochemistry analysis of CD47 KO HEK293T (ab266324) cells labelling CD47 with ab256495 at 1 : 100 (5.62 μg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% TritonX-100. Cells were counterstained with Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) (ab195889) at 1/200 (2.5μg/ml) dilution. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) was used as the secondary antibody at 1 : 1000 (2 ug/ml) dilution. DAPI (blue) was used as nuclear counterstain. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) was used as the secondary antibody only control. Confocal image showing membranous staining in Parental HEK293 cell line, no staining in CD47 KO HEK293T cell line.
- WB
Lab
Western blot - Human CD47 knockout HEK-293T cell line (AB266324)
Lanes 1-4 : Merged signal (red and green). Green - ab218810 observed at 47-52 kDa. Red - loading control ab8245 observed at 36 kDa.
ab218810 Anti-CD47 antibody [EPR21794] was shown to specifically react with CD47 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266324 (knockout cell lysate ab257220) was used. Wild-type and CD47 knockout samples were subjected to SDS-PAGE. ab218810 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4° at 1 in 500 dilution 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.
All lanes:
Western blot - Anti-CD47 antibody [EPR21794] (<a href='/en-us/products/primary-antibodies/cd47-antibody-epr21794-ab218810'>ab218810</a>) at 1/500 dilution
Lane 1:
Wild-type HEK293T cell lysate at 20 µg
Lane 2:
CD47 knockout HEK293T cell lysate at 20 µg
Lane 2:
Western blot - Human CD47 knockout HEK-293T cell line (ab266324)
Lane 3:
Jurkat cell lysate at 20 µg
Lane 4:
HepG2 cell lysate at 20 µg
Secondary
All lanes:
Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution
Predicted band size: 35 kDa
Observed band size: 47-52 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human CD47 knockout HEK-293T cell line (AB266324)
Allele-1 : 11 bp deletion in exon 2
- Cell Culture
Unknown
Cell Culture - Human CD47 knockout HEK-293T cell line (AB266324)
Representative images of CD47 knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T 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.
- Sanger seq
Unknown
Sanger Sequencing - Human CD47 knockout HEK-293T cell line (AB266324)
Allele-2 : 5 bp deletion in exon 2.
- Sanger seq
Lab
Sanger Sequencing - Human CD47 knockout HEK-293T cell line (AB266324)
Sequencing chromatogram displaying sequence edit in exon 2
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Cell Lines & Lysates
AB267231
Human HLA-E (HLA E) knockout HEK-293T cell line
Advanced Validation
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Reactivity data
Product details
We will provide viable cells that proliferate on revival.
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.
What's included?
Properties and storage information
Gene name
Gene editing type
Gene editing method
Knockout validation
Shipped at conditions
Appropriate short-term storage conditions
Appropriate long-term storage conditions
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.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
CD47 functions as a "don't eat me" signal inhibiting phagocytosis by binding to SIRPα on macrophages. This protein is also important for cell adhesion and migration and can modulate interactions with integrins. While not a part of a large complex CD47 associates with various cytoskeletal and membrane proteins to maintain cellular architecture and transmission of mechanical forces. Additionally its interaction with thrombospondins influences angiogenesis and nitric oxide signaling.
Pathways
CD47 is extensively involved in the regulation of the immune response and angiogenesis. The interaction between CD47 and SIRPα plays a part in the regulation of macrophage activity impacting the immune signaling pathway. CD47 also interacts with integrins allowing it to contribute to the angiogenesis pathway which is critical in the formation of new blood vessels. These interactions help coordinate complex cellular responses and maintain tissue homeostasis.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
Target data
Publications (1)
Recent publications for all applications. Explore the full list and refine your search
Nature communications 16:1903 PubMed39988725
2025
Applications
Unspecified application
Species
Unspecified reactive species
Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
For licensing inquiries, please contact partnerships@abcam.com