Human CDKN1A knockout MCF7 cell line
- Advanced Validation
- What is this?
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(0 Publication)
- WB
Lab
Western blot - Human CDKN1A knockout MCF7 cell line (AB288200)
Western blot : Anti-CDKN1A antibody [CIP1/823] (ab220206) staining at 1 ug/ml, shown in green; Rabbit Anti-GAPDH antibody [EPR16891] (ab181602) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab220206 was shown to bind specifically to CDKN1A. A band was observed at 21 kDa in wild-type cell lysates with no signal observed at this size in CDKN1A knockout cell lines. To generate this image, wild-type and CDKN1A knockout cell lysates were analysed. CDKN1A knockout cell lines in HCT 116 (ab288187), MCF7 (ab288200) and A549 (ab288213) backgrounds were used. 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-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution.
All lanes:
Western blot - Anti-p21 antibody [CIP1/823] (<a href='/en-us/products/primary-antibodies/p21-antibody-cip1-823-ab220206'>ab220206</a>) at 1 µg/mL
Lane 1:
Wild-type HCT 116 cell lysate at 20 µg
Lane 2:
Western blot - Human CDKN1A knockout HCT116 cell line (<a href='/en-us/products/cell-lines/human-cdkn1a-knockout-hct116-cell-line-ab288187'>ab288187</a>)
Lane 2:
CDKN1A knockout HCT 116 cell lysate at 20 µg
Lane 3:
Wild-type MCF7 cell lysate at 20 µg
Lane 4:
Western blot - Human CDKN1A knockout MCF7 cell line (ab288200)
Lane 4:
CDKN1A knockout MCF7 cell lysate at 20 µg
Lane 5:
Wild-type A549 cell lysate at 20 µg
Lane 6:
Western blot - Human CDKN1A knockout A549 cell line (<a href='/en-us/products/cell-lines/human-cdkn1a-knockout-a549-cell-line-ab288213'>ab288213</a>)
Lane 6:
CDKN1A knockout A549 cell lysate at 20 µg
Secondary
All lanes:
Goat anti-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution
Predicted band size: 18 kDa
Observed band size: 21 kDa
false
- WB
Lab
Western blot - Human CDKN1A knockout MCF7 cell line (AB288200)
Western blot : Anti-CDKN1A antibody [EPR362] (ab109520) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab109520 was shown to bind specifically to CDKN1A. A band was observed at 21 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in CDKN1A knockout cell line. To generate this image, wild-type and CDKN1A knockout HCT 116 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-p21 antibody [EPR362] (<a href='/en-us/products/primary-antibodies/p21-antibody-epr362-ab109520'>ab109520</a>) at 1/1000 dilution
Lane 1:
Wild-type HCT 116 cell lysate at 20 µg
Lane 2:
Western blot - Human CDKN1A knockout HCT116 cell line (<a href='/en-us/products/cell-lines/human-cdkn1a-knockout-hct116-cell-line-ab288187'>ab288187</a>)
Lane 2:
CDKN1A knockout HCT 116 cell lysate at 20 µg
Lane 3:
Wild-type MCF7 cell lysate at 20 µg
Lane 4:
Western blot - Human CDKN1A knockout MCF7 cell line (ab288200)
Lane 4:
CDKN1A knockout MCF7 cell lysate at 20 µg
Lane 5:
Wild-type A549 cell lysate at 20 µg
Lane 6:
Western blot - Human CDKN1A knockout A549 cell line (<a href='/en-us/products/cell-lines/human-cdkn1a-knockout-a549-cell-line-ab288213'>ab288213</a>)
Lane 6:
CDKN1A knockout A549 cell lysate at 20 µg
Secondary
All lanes:
Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution
Predicted band size: 21 kDa
Observed band size: 21 kDa
false
- NGS
Supplier Data
Next Generation Sequencing - Human CDKN1A knockout MCF7 cell line (AB288200)
221 bp deletion after Gly123 of the WT protein
- Sanger seq
Project
Sanger Sequencing - Human CDKN1A knockout MCF7 cell line (AB288200)
221bp deletion after Ala 5 of the WT protein.
Complementary Products
Primary Antibodies
AB109520
RabMAb|Recombinant|KO Validated|Lab Essentials|20ul selling size
![Western blot - Anti-p21 antibody [EPR362] (AB109520)](https://content.abcam.com/products/images/p21-antibody-epr362-ab109520--western-blot-img422025.jpg)
- 4
- 16
Primary Antibodies
AB108349
Anti-CDKN2A/p16INK4a antibody [EPR1473] - C-terminal
BOND RX™ Validated|20ul selling size|KO Validated|RabMAb|Recombinant
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CDKN2A/p16INK4a antibody [EPR1473] - C-terminal (AB108349)](https://content.abcam.com/products/images/cdkn2a-p16ink4a-antibody-epr1473-c-terminal-ab108349--immunohistochemistry-formalin-pfa-fixed-paraffin-embedded-sections-img74532.jpg)
- 4
- 7
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
- Slow to trypsinise.
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 5-7x104 cells/cm2 is recommended.
- Cells should be passaged when they have achieved 80-90% confluence.
Culture medium
MEM + 10% FBS + 0.01 mg/ml bovine insulin
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
P21 plays a critical role in cell cycle regulation and DNA damage response. It associates with the p53 tumor suppressor protein forming an essential part of the p53 signaling pathway under stress conditions. p21 arrests cells in G1 phase allowing DNA repair or leading to cellular senescence. Through these mechanisms p21 integrates signals from stress and damage emphasizing its importance as a mediator in cellular checkpoint control.
Pathways
P21 is closely involved in the p53 and the Transforming Growth Factor-beta (TGF-β) signaling pathways. This integration helps in transmitting growth-inhibitory signals. p21 interacts with proteins such as p53 and cyclins ensuring precise regulation of the cell cycle and growth arrest when necessary. Triggered by p53 activation p21 contributes to preventing undisturbed cell proliferation acting as a safeguard against tumor formation.
Cell culture
Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Female
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