VIM KO cell line available to order. KO validated by Next Generation Sequencing, Western blot. Free of charge wild type control provided.
Images
Key facts
- Cell type
- A549
- Species or organism
- Human
- Tissue
- Lung
- Form
- Liquid
- Knockout validation
- Next Generation Sequencing, Western blot
Alternative names
CTRCT30, Epididymis luminal protein 113, FLJ36605, HEL113, VIME_HUMAN, Vimentin
Recommended products
VIM KO cell line available to order. KO validated by Next Generation Sequencing, Western blot. Free of charge wild type control provided.
Key facts
- Cell type
- A549
- Species or organism
- Human
- Tissue
- Lung
- Form
- Liquid
- Knockout validation
- Next Generation Sequencing, Western blot
- Disease
- Carcinoma
- Concentration
Properties
- Gene name
- VIM
- Gene editing type
- Knockout
- Gene editing method
- CRISPR technology
- Knockout validation
- Next Generation Sequencing, Western blot
Quality control
- STR analysis
- CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
- Biosafety level
- EU: 1 US: 1
- Adherent/suspension
- Adherent
- Gender
- Male
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.
Storage
- Shipped at conditions
- Dry Ice
- Appropriate short-term storage conditions
- -196°C
- Appropriate long-term storage conditions
- -196°C
Notes
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.
Recommended control: Human wild-type A549 cell line (ab288558). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.
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.
Product promise
We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.
In the unlikely event of one of our products not working as expected, you are covered by our product promise.
Full details and terms and conditions can be found here:
Terms & Conditions.
3 product images
Next Generation Sequencing - Human VIM knockout A549 cell line (ab288984)
127 bp deletion, 17 bp deletion downstream (Allele 1); 127 bp deletion, 3 bp deletion downstream (Allele 2); 127 bp deletion; 222 bp insertion downstream (Allele 3)
Western blot - Human VIM knockout A549 cell line (ab288984)
This image was generated using a previous batch manufactured using the hybridoma production method.
False colour image of Western blot: Anti-Vimentin antibody [SP20] staining at 1/120 dilution, shown in green; Mouse anti-CANX [CANX/1543] (Anti-Calnexin antibody [CANX/1543] ab238078) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-Vimentin antibody [SP20] ab16700 was shown to bind specifically to Vimentin. A band was observed at 55 kDa in wild-type A549 cell lysates with no signal observed at this size in VIM knockout cell line ab288984. To generate this image, wild-type and VIM 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 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
All lanes: Western blot - Anti-Vimentin antibody [SP20] (Anti-Vimentin antibody [SP20] ab16700) at 1/120 dilution
Lane 1: Wild-type A549 cell lysate at 20 µg
Lane 2: Vimentin knockout A549 cell lysate at 20 µg
Lane 3: Daudi cell lysate at 20 µg
Performed under reducing conditions.
Observed band size: 55 kDa
Western blot - Human VIM knockout A549 cell line (ab288984)
False colour image of Western blot: Anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (Anti-Calnexin antibody [CANX/1543] ab238078) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker ab92547 was shown to bind specifically to Vimentin. A band was observed at 55 kDa in wild-type A549 cell lysates with no signal observed at this size in VIM knockout cell line ab288984. To generate this image, wild-type and VIM 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 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
All lanes: Western blot - Anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker (Anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker ab92547) at 1/1000 dilution
Lane 1: Wild-type A549 cell lysate at 20 µg
Lane 2: VIM knockout A549 cell lysate at 20 µg
Lane 3: Daudi cell lysate at 20 µg
Performed under reducing conditions.
Observed band size: 55 kDa
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