Human EGFR knockout A549 cell line
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- WB
Lab
Western blot - Human EGFR knockout A549 cell line (AB286394)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52894).
Western blot : Anti-EGFR antibody [EP38Y] (ab52894) staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab52894 was shown to bind specifically to EGFR. A band was observed at 175 kDa in wild-type A549 cell lysates with no signal observed at this size in EGFR knockout cell line. To generate this image, wild-type and EGFR knockout A549 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 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.
All lanes:
Western blot - Anti-EGFR antibody [EP38Y] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/egfr-antibody-ep38y-bsa-and-azide-free-ab272293'>ab272293</a>)
Lane 1:
Wild-type A549 cell lysate at 20 µg
Lane 2:
EGFR knockout A549 cell lysate at 20 µg
Lane 3:
Wild-type HeLa cell lysate at 20 µg
Lane 4:
EGFR knockout HeLa cell lysate at 20 µg
Observed band size: 175 kDa
true
- WB
Lab
Western blot - Human EGFR knockout A549 cell line (AB286394)
Western blot : Anti-EGFR antibody [E234] (ab32198) 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, ab32198 was shown to bind specifically to EGFR. A band was observed at 160 kDa in wild-type cell lysates with no signal observed at this size in EGFR knockout cell lines. To generate this image, wild-type and EGFR knockout A549 (ab286394) and HeLa (ab255385) 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.
All lanes:
Western blot - Anti-EGFR antibody [E234] (<a href='/en-us/products/primary-antibodies/egfr-antibody-e234-ab32198'>ab32198</a>) at 1/1000 dilution
Lane 1:
Wild-type A549 cell lysate at 20 µg
Lane 2:
Western blot - Human EGFR knockout A549 cell line (ab286394)
Lane 2:
EGFR knockout A549 cell lysate at 20 µg
Lanes 2 and 4:
Western blot - Human EGFR knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-egfr-knockout-hela-cell-line-ab255385'>ab255385</a>)
Lane 3:
Wild-type HeLa cell lysate at 20 µg
Lane 4:
EGFR knockout HeLa cell lysate at 20 µg
Observed band size: 160 kDa
false
- WB
Lab
Western blot - Human EGFR knockout A549 cell line (AB286394)
Western blot : Anti-EGFR antibody [E235] (ab32077) 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, ab32077 was shown to bind specifically to EGFR. A band was observed at 160 kDa in wild-type cell lysates with no signal observed at this size in EGFR knockout cell lines. To generate this image, wild-type and EGFR knockout A549 (ab286394) and HeLa (ab255385) 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.
All lanes:
Western blot - Anti-EGFR antibody [E235] (<a href='/en-us/products/primary-antibodies/egfr-antibody-e235-ab32077'>ab32077</a>) at 1/1000 dilution
Lane 1:
Wild-type A549 cell lysate at 20 µg
Lane 2:
Western blot - Human EGFR knockout A549 cell line (ab286394)
Lane 2:
EGFR knockout A549 cell lysate at 20 µg
Lanes 2 and 4:
Western blot - Human EGFR knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-egfr-knockout-hela-cell-line-ab255385'>ab255385</a>)
Lane 3:
Wild-type HeLa cell lysate at 20 µg
Lane 4:
EGFR knockout HeLa cell lysate at 20 µg
Observed band size: 160 kDa
false
- Sanger seq
Supplier Data
Sanger Sequencing - Human EGFR knockout A549 cell line (AB286394)
2 bp deletion after the Gln145 and 8 bp deletion after the Glu160 of the WT protein
- NGS
Supplier Data
Next Generation Sequencing - Human EGFR knockout A549 cell line (AB286394)
6 bp (allele 1), 3 bp (allele 2), and 10 bp deletion after Ser160 of the WT protein
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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.
- Do not allow the cell density to exceed 7x104 cells/cm2.
Culture medium
F-12K + 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
The EGFR protein plays an important role in cellular communication and signaling processes. EGFR pairs with other receptor family members to form active dimers or even higher-order complexes which in turn initiate intracellular signaling cascades. Through these complexes EGFR influences many processes including cell differentiation and repair. This function of EGFR makes it an integral part of mammalian biology affecting how cells respond to their environment by mediating changes in gene expression.
Pathways
EGFR is a central player in the MAPK and PI3K/Akt signaling pathways. Alongside other protein partners like KRAS and PI3 kinase it contributes to transmitting signals from the cell surface to the nucleus affecting gene transcription and cell behavior. These pathways are important for normal cell growth and division and aberrations in these pathways can lead to excessive or insufficient cell proliferation.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Male
Viability
~ 60%
Target data
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