Human MICA knockout A549 cell line
- Advanced Validation
- What is this?
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- WB
Lab
Western blot - Human MICA knockout A549 cell line (AB288893)
Western blot : Anti-MICA antibody [EPR22031] (ab224702) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab224702 was shown to bind specifically to MICA. A band was observed at 43 kDa in wild-type A549 cell lysates with no signal observed at this size in MICA knockout cell line. To generate this image, wild-type and MICA 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-MICA + MICB antibody [EPR22031] (<a href='/en-us/products/primary-antibodies/mica-micb-antibody-epr22031-ab224702'>ab224702</a>) at 1/1000 dilution
Lane 1:
Wild-type A549 cell lysate at 20 µg
Lane 2:
MICA knockout A549 cell lysate at 20 µg
Lane 3:
U-2 OS cell lysate at 20 µg
Lane 4:
Ramos cell lysate at 20 µg
Lane 5:
MICA protein <a href='/en-us/products/proteins-peptides/recombinant-human-mica-protein-active-ab182709'>ab182709</a> cell lysate at 0.2 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: 43 kDa
false
- NGS
Lab
Next Generation Sequencing - Human MICA knockout A549 cell line (AB288893)
94 bp deletion after Met 207 (allele 1); 2 bp deletion after Ala 228 (allele 2) of WT protein
Complementary Products
Primary Antibodies
AB224702
Anti-MICA + MICB antibody [EPR22031]
KO Validated|RabMAb|Recombinant
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MICA + MICB antibody [EPR22031] (AB224702)](https://content.abcam.com/products/images/mica-micb-antibody-epr22031-ab224702--immunohistochemistry-formalin-pfa-fixed-paraffin-embedded-sections-img10757.jpg)
- 0
- 0
![Flow Cytometry - Anti-TCR beta F1 antibody [8A3] (AB171088)](https://content.abcam.com/products/images/tcr-beta-f1-antibody-8a3-ab171088--flow-cytometry-img81288.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
The MICA protein acts as a ligand for the NKG2D receptor an activating receptor found on NK cells and some T-cell subsets. MICA does not require expression as part of a complex with β2-microglobulin for function. When cells express MICA they signal distress allowing immune cells to target and destroy potentially harmful cells such as those infected with viruses or transformed into cancer cells. This interaction is an important component of the immune surveillance system.
Pathways
MICA functions within the natural cytotoxicity effector pathway heavily involved in immune responses. It is also relevant in the DNA damage response pathways. Within these pathways MICA interacts with proteins like NKG2D and plays a part in mediating cellular responses to stress and damage. These interactions enhance the capacity of immune cells to recognize and eliminate abnormal cells.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Male
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