TP53 KO cell line available to order. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 49 bp deletion in exon 4.
Antigen NY-CO-13, BCC7, Cellular tumor antigen p53, FLJ92943, LFS1, Mutant tumor protein 53, P53_HUMAN, Phosphoprotein p53, TRP53, Tp53, Transformation related protein 53, Tumor protein 53, Tumor protein p53, Tumor suppressor p53, p53 tumor suppressor, tumor antigen p55
TP53 KO cell line available to order. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 49 bp deletion in exon 4.
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.
We will provide viable cells that proliferate on revival.
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False colour image of Western blot: Anti-p53 antibody [E26] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-p53 antibody [E26] ab32389 was shown to bind specifically to p53. A band was observed at 49 kDa in wild-type A549 cell lysates with no signal observed at this size in tp53 knockout cell line ab276092 (knockout cell lysate ab282999). To generate this image, wild-type and tp53 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 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 (IRDye® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/20000 dilution.
All lanes: Western blot - Anti-p53 antibody [E26] (Anti-p53 antibody [E26] ab32389) at 1/1000 dilution
Lane 1: Wild-type A549 cell lysate at 20 µg
Lane 2: TP53 knockout A549 cell lysate at 20 µg
Lane 2: Western blot - Human TP53 (p53) knockout Jurkat cell line (Human TP53 (p53) knockout Jurkat cell line ab276112)
Lane 3: Wild-type Jurkat cell lysate at 20 µg
Lane 4: TP53 knockout Jurkat cell lysate at 20 µg
Lane 5: A431 cell lysate at 20 µg
Lane 6: Saos-2 cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 43 kDa
Observed band size: 49 kDa
False colour image of Western blot: Anti-p53 antibody [DO-1] - ChIP Grade staining at 1/1000 dilution, shown in green; Rabbit anti-alpha Tubulin antibody [EP1332Y] (Anti-alpha Tubulin antibody [EP1332Y] - Loading Control ab52866) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-p53 antibody [DO-1] - ChIP Grade ab1101 was shown to bind specifically to p53. A band was observed at 49 kDa in wild-type A549 cell lysates with no signal observed at this size in tp53 knockout cell line ab276092 (knockout cell lysate ab282999). To generate this image, wild-type and tp53 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 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 (IRDye® 800CW) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed ab216777) at 1/20000 dilution.
All lanes: Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (Anti-p53 antibody [DO-1] - ChIP Grade ab1101) at 1/1000 dilution
Lane 1: Wild-type A549 cell lysate at 20 µg
Lane 2: Western blot - Human TP53 (p53) knockout A549 cell line (ab276092) at 20 µg
Lane 3: Wild-type Jurkat cell lysate at 20 µg
Lane 4: Western blot - Human TP53 (p53) knockout Jurkat cell line (Human TP53 (p53) knockout Jurkat cell line ab276112) at 20 µg
Lane 5: A431 cell lysate at 20 µg
Lane 6: Saos-2 cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 43 kDa
Observed band size: 49 kDa
49 bp deletion in exon 4
False colour image of Western blot: Anti-p53 antibody [DO-1] - ChIP Grade staining at 1/1000 dilution, shown in green; Rabbit Anti-GAPDH antibody [EPR16891] (Anti-GAPDH antibody [EPR16891] - Loading Control ab181602) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-p53 antibody [DO-1] - ChIP Grade ab1101 was shown to bind specifically to p53. A band was observed at 40/50 kDa in treated wild-type A549 cell lysates with no signal observed at this size in tp53 knockout cell line ab276092. To generate this image, wild-type and tp53 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 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 (IRDye® 800CW) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed ab216777) at 1/20000 dilution.
Lanes 1 - 12: Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (Anti-p53 antibody [DO-1] - ChIP Grade ab1101) at 1/1000 dilution
Lanes 1 - 12: Western blot - Anti-p53 antibody [DO-1] - BSA and Azide free (Anti-p53 antibody [DO-1] - BSA and Azide free ab237976)
Lane 1: MCF7 Treated Camptothecin (20 ug/ml, 0 h) cell lysate at 15 µg
Lane 2: MCF7 Treated Camptothecin (20 ug/ml, 6 h) cell lysate at 15 µg
Lane 3: MCF7 Treated Camptothecin (20 ug/ml, 24 h) cell lysate at 15 µg
Lane 4: Wild-type A549 Treated Camptothecin (20 ug/ml, 0 h) cell lysate at 15 µg
Lane 5: Wild-type A549 Treated Camptothecin (20 ug/ml, 6 h) cell lysate at 15 µg
Lane 6: Wild-type A549 Treated Camptothecin (20 ug/ml, 24 h) cell lysate at 15 µg
Lane 7: TP53 knockout A549 Treated Camptothecin (20 ug/ml, 0 h) cell lysate at 15 µg
Lane 8: TP53 knockout A549 Treated Camptothecin (20 ug/ml, 6 h) cell lysate at 15 µg
Lane 9: TP53 knockout A549 Treated Camptothecin (20 ug/ml, 24 h) cell lysate at 15 µg
Lane 10: Saos-2 cell lysate at 15 µg
Lane 11: SK-BR-3 cell lysate at 15 µg
Lane 12: A431 cell lysate at 15 µg
All lanes: Western blot - Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed ab216772) at 1/20000 dilution
Performed under reducing conditions.
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