Human FADD knockout HeLa cell line
- Advanced Validation
- What is this?
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FADD KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: Insertion of the selection cassette in exon 1.
View Alternative Names
FADD protein, FADD_HUMAN, FAS-associated death domain protein, FAS-associating death domain-containing protein, Fas (TNFRSF6) associated via death domain, Fas TNFRSF6 associated via death domain, Fas associated via death domain, Fas associating protein, Fas associating protein with death domain, GIG 3, Growth-inhibiting gene 3 protein, H sapiens mRNA for mediator of receptor induced toxicity, MGC8528, MORT 1, Mediator of receptor induced toxicity, Protein FADD
- WB
Lab
Western blot - Human FADD knockout HeLa cell line (AB261817)
Lanes 1 - 2 : Merged signal (red and green). Green - ab108601 observed at 23 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.
ab108601 was shown to react with FADD in wild-type HeLa cells in western blot with loss of signal observed in FADD knockout cell line ab261817 (FADD knockout cell lysate ab257261). Wild-type and FADD knockout HeLa cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab108601 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-FADD antibody [EPR4415] (<a href='/en-us/products/primary-antibodies/fadd-antibody-epr4415-ab108601'>ab108601</a>) at 1/1000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
FADD knockout HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human FADD knockout HeLa cell line (ab261817)
Predicted band size: 23 kDa
Observed band size: 23 kDa
false
- WB
Lab
Western blot - Human FADD knockout HeLa cell line (AB261817)
Lanes 1-4 : Merged signal (red and green). Green - ab119059 observed at 25 kDa. Red - loading control ab181602 observed at 37 kDa.
ab119059 Anti-FADD antibody [OTI1C11] was shown to specifically react with FADD in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab261817 (knockout cell lysate ab257261) was used. Wild-type and FADD knockout samples were subjected to SDS-PAGE. ab119059 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) were incubated overnight at 4°C at 1 in 2000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-FADD antibody [OTI1C11] (<a href='/en-us/products/primary-antibodies/fadd-antibody-oti1c11-ab119059'>ab119059</a>) at 1/2000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
FADD knockout HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human FADD knockout HeLa cell line (ab261817)
Lane 3:
A431 cell lysate at 20 µg
Lane 4:
Jurkat cell lysate at 20 µg
Secondary
All lanes:
Western blot - Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-800cw-preadsorbed-ab216772'>ab216772</a>) at 1/20000 dilution
Predicted band size: 23 kDa
Observed band size: 25 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human FADD knockout HeLa cell line (AB261817)
Homozygous : Insertion of the selection cassette in exon 1.
Reactivity data
Product details
Recommended control: Human wild-type HeLa cell line (ab255928). 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, 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.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
FADD is essential in apoptosis where it assists in the assembly of the death-inducing signaling complex (DISC). Upon receptor activation FADD recruits procaspase-8 or -10 to DISC promoting their autocatalytic cleavage and activation. This leads to the subsequent cascade that results in cell apoptosis. FADD also plays a role in necroptosis and is involved in the immune response regulation highlighting its multifunctional nature in cellular processes.
Pathways
FADD integrates into the apoptotic and necroptotic pathways. In the apoptotic pathway it interacts closely with Fas a death receptor to promote caspase-8 activation. Additionally in the necroptotic pathway FADD associates with RIP1 and RIP3 contributing to an alternative form of programmed cell death. These interactions underline its significant role in controlling cell fate decisions.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
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Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
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