Human DYRK1A knockout HEK-293T cell line
- Advanced Validation
- What is this?
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- WB
Lab
Western blot - Human DYRK1A knockout HEK-293T cell line (AB267336)
False colour image of Western blot : Anti-DYRK1A antibody [EPR24132-61] staining at 1/2000 dilution shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution shown in red. In Western blot ab259869 was shown to bind specifically to DYRK1A. A band was observed at 80/85/90 kDa in wild-type HEK-293T cell lysates with no signal observed at this size in DYRK1A CRISPR-Cas9 edited cell line ab267336 (CRISPR-Cas9 edited cell lysate ab258402). The band observed in the CRISPR-Cas9 edited lysate lane below 80/85/90 kDa is likely to represent a truncated form of DYRK1A. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image wild-type and DYRK1A CRISPR-Cas9 edited HEK-293T 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 °. 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 (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
All lanes:
Western blot - Anti-DYRK1A antibody [EPR24132-61] (<a href='/en-us/products/primary-antibodies/dyrk1a-antibody-epr24132-61-ab259869'>ab259869</a>) at 1/2000 dilution
Lane 1:
Wild-type HEK-293T cell lysate at 20 µg
Lane 2:
DYRK1A CRISPR-Cas9 edited HEK-293T cell lysate at 20 µg
Lane 2:
Western blot - Human DYRK1A knockout HEK-293T cell line (ab267336)
Lane 3:
HT1080 cell lysate at 20 µg
Lane 4:
U-251 MG cell lysate at 20 µg
Secondary
All lanes:
Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution
Predicted band size: 86 kDa
Observed band size: 80 kDa,85 kDa,90 kDa
false
- WB
Lab
Western blot - Human DYRK1A knockout HEK-293T cell line (AB267336)
False colour image of Western blot : Anti-DYRK1A antibody staining at 1/500 dilution, shown in green; Rabbit Anti-GAPDH antibody [EPR16891] (ab181602) loading control staining at 1/20000 dilution, shown in red. In Western blot, the antibody was shown to bind specifically to DYRK1A. A band was observed at 80/95/90 kDa in wild-type HEK-293T cell lysates with no signal observed at this size in DYRK1A knockout cell line ab267336 (knockout cell lysate ab258402). To generate this image, wild-type and DYRK1A knockout HEK-293T 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 (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) at 1/20000 dilution.
All lanes:
Anti-DYRK1A antibody at 1/500 dilution
Lane 1:
Wild-type HEK-293T cell lysate at 20 µg
Lane 2:
HEK-293T cell lysate at 20 µg
Lane 2:
Western blot - Human DYRK1A knockout HEK-293T cell line (ab267336)
Lane 3:
HT1080 cell lysate at 20 µg
false
- Sanger seq
Unknown
Sanger Sequencing - Human DYRK1A knockout HEK-293T cell line (AB267336)
Homozygous : 1 bp insertion in exon12
Complementary Products
Primary Antibodies
AB259869
Anti-DYRK1A antibody [EPR24132-61]
20ul selling size|RabMAb|Recombinant
![Western blot - Anti-DYRK1A antibody [EPR24132-61] (AB259869)](https://content.abcam.com/products/images/dyrk1a-antibody-epr24132-61-ab259869--western-blot-img174742.jpg)
- 0
- 0
Reactivity data
Product details
We will provide viable cells that proliferate on revival.
Western blot data indicates that the CRISPR gene edit may have resulted in a truncation of the protein of interest. Please see data images.
This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and 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
Zygosity
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.
Culture medium
DMEM (High Glucose) + 10% FBS
Cryopreservation medium
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
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