Human IRF5 knockout A549 cell line
- Advanced Validation
- What is this?
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- WB
Lab
Western blot - Human IRF5 knockout A549 cell line (AB301006)
Western blot : Anti-IRF5 antibody [EPR17067] (ab181553) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab181553 was shown to bind specifically to IRF5. A band was observed at 50-70 kDa in wild-type A549 cell lysates with no signal observed at this size in IRF5 knockout cell line. To generate this image, wild-type and IRF5 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-IRF5 antibody [EPR17067] (<a href='/en-us/products/primary-antibodies/irf5-antibody-epr17067-ab181553'>ab181553</a>) at 1/1000 dilution
Lane 1:
Wild-type A549 cell lysate at 20 µg
Lane 2:
IRF5 knockout A549 cell lysate at 20 µg
Secondary
All lanes:
Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution
Observed band size: 50 kDa,70 kDa
false
- WB
Lab
Western blot - Human IRF5 knockout A549 cell line (AB301006)
Western blot : Anti-CSK antibody [EPR24673-97] (ab300132) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab300132 was shown to bind specifically to CSK. A band was observed at 50 kDa in wild-type A549 cell lysates with no signal observed at this size in CSK knockout cell line. To generate this image, wild-type and CSK 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-IRF5 antibody [10T1] (<a href='/en-us/products/primary-antibodies/irf5-antibody-10t1-ab33478'>ab33478</a>) at 1/1000 dilution
Lane 1:
Wild-type A549 cell lysate at 20 µg
Lane 2:
IRF5 knockout A549 cell lysate at 20 µg
Lane 2:
Western blot - Human IRF5 knockout A549 cell line (ab301006)
Secondary
All lanes:
Goat anti-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution
Observed band size: 63 kDa
false
- NGS
Lab
Next Generation Sequencing - Human IRF5 knockout A549 cell line (AB301006)
113 bp deletion after Pro 82 (allele 1); 59 bp deletion after Ala 83 and 14 bp deletion after Gly 99 (allele 2) of WT protein
Complementary Products
![Western blot - Anti-IRF5 antibody [EPR17067] (AB181553)](https://content.abcam.com/products/images/irf5-antibody-epr17067-ab181553--western-blot-img3959.jpg)
Reactivity data
Product details
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.
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.
- 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.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
IRF5 influences the immune response through the regulation of cytokine production. It activates genes involved in producing pro-inflammatory cytokines such as TNF-? and IL-6 which are important for clearing viral infections and activating immune cells. IRF5 does not usually form part of a protein complex but it interacts closely with TRAF6 and MyD88 for its activation. Its pivotal role in cytokine production highlights its importance in controlling inflammation.
Pathways
IRF5 is an important component of the Toll-like receptor (TLR) signaling pathway and plays a role in the MyD88-dependent pathway. In this context IRF5 gets activated by TLRs interacting with MyD88 and TAK1 to enhance the transcription of important genes for immune response. It is also closely connected to the IRF3 protein which shares similar activation pathways and can similarly initiate an antiviral state.
Cell culture
Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Male
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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