Human E2F4 knockout HEK-293T cell line
- Advanced Validation
- What is this?
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(0 Publication)
- WB
Lab
Western blot - Human E2F4 knockout HEK-293T cell line (AB266119)
Lanes 1-4 : Merged signal (red and green). Green - ab150360 observed at 60 kDa. Red - loading control ab8245 observed at 36 kDa.
ab150360 Anti-E2F4 antibody [EPR8259] was shown to specifically react with E2F4 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266119 (knockout cell lysate ab257932) was used. Wild-type and E2F4 knockout samples were subjected to SDS-PAGE. ab150360 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2.5 hours at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-E2F4 antibody [EPR8259] (<a href='/en-us/products/primary-antibodies/e2f4-antibody-epr8259-ab150360'>ab150360</a>) at 1/1000 dilution
Lane 1:
Wild-type HEK293T cell lysate at 20 µg
Lane 2:
E2F4 knockout HEK293T cell lysate at 20 µg
Lane 2:
Western blot - Human E2F4 knockout HEK-293T cell line (ab266119)
Lane 3:
K-562 cell lysate at 20 µg
Lane 4:
Jurkat cell lysate at 20 µg
Secondary
All lanes:
Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution
Predicted band size: 44 kDa
Observed band size: 60 kDa
false
- Sanger seq
Lab
Sanger Sequencing - Human E2F4 knockout HEK-293T cell line (AB266119)
Allele-2 : Insertion of the selection cassette in exon 2.
- Sanger seq
Unknown
Sanger Sequencing - Human E2F4 knockout HEK-293T cell line (AB266119)
Allele-1 : 4 bp deletion in exon2
- Cell Culture
Lab
Cell Culture - Human E2F4 knockout HEK-293T cell line (AB266119)
Representative images E2F4 knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using a EVOS M5000 microscope.
Complementary Products
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-E2F4 antibody [EPR8259] (AB150360)](https://content.abcam.com/products/images/e2f4-antibody-epr8259-ab150360--immunohistochemistry-formalin-pfa-fixed-paraffin-embedded-sections-img138282.jpg)
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, 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
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.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
E2F4 functions as a regulator of cell cycle progression. It forms complexes localizing mostly in the nucleus controlling the transition from G1 to S phase by regulating genes associated with cell cycle arrest and DNA replication. Partnering with members of the DP family enhances its DNA binding capacity influencing genes involved in cell cycle arrest and DNA repair processes. E2F4 predominantly acts as a transcriptional repressor in quiescent and differentiating cells maintaining the cell in a non-proliferative state.
Pathways
E2F4 is significant in the regulation of the cell cycle and DNA damage response pathways. It engages with the retinoblastoma protein family (pRB p107 p130) to exert repressive effects on E2F-target genes ensuring proper cell cycle exit and maintaining the G0 phase. The interaction with p130 is instrumental during the G0 state preventing inappropriate cell proliferation and contributing to the maintenance of cellular quiescence.
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'.
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