Human PTEN knockout VCaP cell line
Human PTEN knockout VCaP cell line
- KO Validated
- Advanced Validation
- 了解详情
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(0 Publication)
- NGS
Lab
Next Generation Sequencing - Human PTEN knockout VCaP cell line (AB326417)
133 bp deletion (allele 1) and 134 bp deletion (allele 2) after Pro96 in exon 5 (CCDS31238.1).
- WB
Lab
Western blot - Human PTEN knockout VCaP cell line (AB326417)
False colour image of Western blot : Anti-PTEN antibody [Y184] (ab32199) staining at 1/10000 dilution, shown in green; Mouse anti-GAPDH loading control staining, shown in magenta.
In Western blot, ab32199 was shown to bind specifically to PTEN. A band was observed at 47 kDa in Wild-type VCAP cell lysates with no signal observed at this size in PTEN knockout cell line.
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) preadsorbed at 1 : 10000 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed at 1 : 10000.
All lanes:
Western blot - Anti-PTEN antibody [Y184] (<a href='/products/primary-antibodies/pten-antibody-y184-ab32199'>ab32199</a>) at 1/10000 dilution
Lane 1:
Wild-type VCAP cell lysates
Lane 2:
Western blot - Human PTEN knockout VCaP cell line (ab326417)
Secondary
All lanes:
Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed at 1/10000 dilution
Observed band size: 47 kDa
false
- WB
Lab
Western blot - Human PTEN knockout VCaP cell line (AB326417)
False colour image of Western blot : Anti-PTEN antibody [EPR9941] (ab154812) staining at 1/10000 dilution, shown in green; Mouse anti-GAPDH loading control staining, shown in magenta.
In Western blot, ab154812 was shown to bind specifically to PTEN. A band was observed at 47 kDa in Wild-type VCAP cell lysates with no signal observed at this size in PTEN knockout cell line.
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) preadsorbed at 1 : 10000 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed at 1 : 10000.
All lanes:
Western blot - Anti-PTEN antibody [EPR9941] (<a href='/products/primary-antibodies/pten-antibody-epr9941-ab154812'>ab154812</a>) at 1/10000 dilution
Lane 1:
Wild-type VCAP cell lysates
Lane 2:
Western blot - Human PTEN knockout VCaP cell line (ab326417)
Secondary
All lanes:
Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed at 1/10000 dilution
Observed band size: 47 kDa
false
反应性数据
产品详情
Although we aim to provide customers with a homozygous clone, feasibility will be dependent on the biology of the protein. Should only heterozygous edits be achieved, you will be notified of the outcome and be asked to confirm whether the cell line is acceptable. All clones will be accompanied with DNA sequencing data, and the mutation description.
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.
规格
性能和储存信息
基因名称
基因编辑类型
基因编辑方法
敲除验证
运输条件
推荐的短期储存条件
推荐的长期储存条件
处理步骤
初始处理指南
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 280 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% CO.
VCaP is a very slow growing cell line and can take up to 48 hours to attach post-thaw and after subcultures. lt can routinely take a minimum of 2 weeks or more for cells to reach approximately 50% confluence with a dense mixture of adherent cells, floating clusters and moderate to heavy debris is always present. The cells may recover better in a T-25 flask compared to a T75 flask. Do not discard any floating cells that may be present during medium changes and subcultures, instead, spin them down using gentle centrifugation and add them back to the adherent population. The cells attach in small tightly formed clusters and some single cells. As the cells attach and start to proliferate and spread, they will grow as flattened epithelial like islands of tightly packed cells.
传代培养指南
- Subculture when cell density reaches 1-2x105 cells/mL.
- A guide seeding density of 2-4x10e4 cells/mL is recommended.
- All seeding densities should be based on cell counts gained by established methods.
培养基
DMEM + 10% FBS
低温储藏试剂
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
细胞培养
生物安全等级
EU: 2 US: 2
贴壁/悬浮
Adherent
性别
Male
活率
~ 60%
Abcam Product Promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
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