SEMA4D KO cell line available to order. KO validated by Next Generation Sequencing, Western blot. Free of charge wild type control provided.
Images
Key facts
- Cell type
- A549
- Species or organism
- Human
- Tissue
- Lung
- Form
- Liquid
- Knockout validation
- Next Generation Sequencing, Western blot
Alternative names
A8, BB 18, CD 100, CD100 antigen, Coll 4, Collapsin 4, GR3, Leukocyte activation antigen CD100, M sema G, MSEMA, SEM4D_HUMAN, SEMA J, Sema 4d, Sema H, Sema domain immunoglobulin domain Ig transmembrane domain TM and short cytoplasmic domain semaphorin 4D, Semacl 2, Semaphorin C like 2, Semaphorin H, Semaphorin J, Semaphorin-4D, Semcl 2
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SEMA4D KO cell line available to order. KO validated by Next Generation Sequencing, Western blot. Free of charge wild type control provided.
Key facts
- Cell type
- A549
- Species or organism
- Human
- Tissue
- Lung
- Form
- Liquid
- Knockout validation
- Next Generation Sequencing, Western blot
- Disease
- Carcinoma
- Concentration
Properties
- Gene name
- SEMA4D
- Gene editing type
- Knockout
- Gene editing method
- CRISPR technology
- Knockout validation
- Next Generation Sequencing, Western blot
Quality control
- STR analysis
- CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
- Biosafety level
- EU: 1 US: 1
- Adherent/suspension
- Adherent
- Gender
- Male
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.
Storage
- Shipped at conditions
- Dry Ice
- Appropriate short-term storage conditions
- -196°C
- Appropriate long-term storage conditions
- -196°C
Notes
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.
Recommended control: Human wild-type A549 cell line (ab288558). 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 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.
Supplementary info
- This supplementary information is collated from multiple sources and compiled automatically.
- Activity summary
Semaphorin 4D also known as CD100 is a transmembrane protein that plays important roles in cellular signaling. It belongs to the semaphorin protein family known for their role in axonal guidance. CD100 has a molecular mass of approximately 150 kDa. This protein expresses mainly in the immune system including T cells and B cells but can also be found in certain neurons and endothelial cells. It can exist in both a membrane-bound form and a soluble form which arise from proteolytic cleavage.
- Biological function summary
CD100 influences various cellular functions by interacting with multiple receptors such as Plexin-B1 and CD72 as part of a larger signaling complex. It contributes to immune response regulation by promoting T-cell activation and B-cell aggregation which are essential for adaptive immunity. In the nervous system it guides neural development including axon growth and synaptic plasticity. CD100's interaction with Plexin-B1 plays a significant role in these developmental processes often affecting cell morphology and movement.
- Pathways
Semaphorin 4D is involved in both immune and neural signaling pathways. In the immune system it influences the PI3K-Akt signaling pathway which impacts cell survival and immune cell activation. It associates with pathways involving Plexin-B1 and RhoA to regulate cytoskeletal dynamics affecting cell motility and growth. Additionally CD100's role in neural pathways intersects with proteins like neuropilin-1 influencing semaphorin-mediated signaling essential for neural connectivity.
- Associated diseases and disorders
Semaphorin 4D has links to cancer and autoimmune diseases. High expression levels of CD100 are observed in several tumor types where it might promote tumor growth and metastasis through its signaling pathways. In autoimmune conditions like multiple sclerosis CD100 affects immune cell interactions and can exacerbate inflammatory responses. Its involvement with proteins such as CD72 and Plexin-B1 in these conditions highlights its contribution to pathogenesis and potential as a therapeutic target.
Product promise
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Full details and terms and conditions can be found here:
Terms & Conditions.
2 product images
Western blot - Human SEMA4D knockout A549 cell line (ab288912)
Western blot: Anti-SEMA4D antibody [EP3569] (Anti-Semaphorin 4D/CD100 antibody [EP3569] ab134128) staining at 1/2000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, Anti-Semaphorin 4D/CD100 antibody [EP3569] ab134128 was shown to bind specifically to SEMA4D. A band was observed at 95-100 kDa in wild-type A549 cell lysates with no signal observed at this size in SEMA4D knockout cell line. To generate this image, wild-type and SEMA4D 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-Semaphorin 4D/CD100 antibody [EP3569] (Anti-Semaphorin 4D/CD100 antibody [EP3569] ab134128) at 1/2000 dilution
Lane 1: Wild-type A549 cell lysate at 40 µg
Lane 2: SEMA4D knockout A549 cell lysate at 40 µg
Lane 3: HDLM-2 cell lysate at 20 µg
Lane 4: PC-3 cell lysate at 20 µg
SecondaryAll lanes: Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution
Performed under reducing conditions.
Next Generation Sequencing - Human SEMA4D knockout A549 cell line (ab288912)
2 bp deletion and 62 bp deletion after Pro 208 of WT protein
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