Human MCAM (CD146) knockout HeLa cell line
- Advanced Validation
- What is this?
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MCAM KO cell line available to order. KO validated. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, Homozygous: 19 bp deletion in exon 6. To order both knockout and wild-type control cells: select '2 x 1000000 Cells/vial'. To order only knockout cells: select '1000000 Cells/vial'.
View Alternative Names
A32 antigen, CD146 antigen, Cell surface glycoprotein MUC18, Cell surface glycoprotein P1H12, Gicerin, MUC18_HUMAN, Mcam, MelCAM, Melanoma adhesion molecule, Melanoma associated glycoprotein MUC18, Melanoma cell adhesion molecule, Melanoma-associated antigen A32, Melanoma-associated antigen MUC18, S endo 1, S-endo 1 endothelial-associated antigen
- WB
Lab
Western blot - Human MCAM (CD146) knockout HeLa cell line (AB261790)
Lanes 1- 2 : Merged signal (red and green). Green - ab75769 observed at 120 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab75769 was shown to react with CD146 in wild-type HeLa cells in western blot. The band observed in CRISPR/Cas9 edited cell line ab261790 (CRISPR/Cas9 edited cell lysate ab256985) lane below 120kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Wild-type HeLa and MCAM CRISPR/Cas9 edited HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab75769 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4° at a 1 in 1000 dilution and a 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-CD146 antibody [EPR3208] (<a href='/en-us/products/primary-antibodies/cd146-antibody-epr3208-ab75769'>ab75769</a>) at 1/1000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
MCAM CRISPR/Cas9 edited HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human MCAM (CD146) knockout HeLa cell line (ab261790)
Predicted band size: 72 kDa
Observed band size: 120 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human MCAM (CD146) knockout HeLa cell line (AB261790)
Homozygous : 19 bp deletion in exon 6.
Reactivity data
Product details
We will provide viable cells that proliferate on revival.
Western blot data indicates that the CRISPR gene edit may have resulted in a truncation of the protein of interest. Please see data images.
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
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
CD146 influences the migration and organization of cells within the vascular system. It serves as a component of cell adhesion complexes facilitating interactions between endothelial cells and other cell types. Additionally CD146 contributes to angiogenesis the process by which new blood vessels form from existing vasculature which is critical during tissue development and repair. This function makes it an important element in maintaining normal physiological processes.
Pathways
CD146 participates in the Wnt/β-catenin and MAPK signaling pathways which are vital in cell proliferation and differentiation. Through the Wnt/β-catenin pathway CD146 interacts with other proteins like Frizzled receptors to regulate gene transcription. In the MAPK pathway it is linked with signaling cascades that involve proteins such as ERK leading to cellular responses necessary for growth and response to external stimuli.
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
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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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