Cat.No. : CSC-RO0121 Host Cell: Ba/F3
Size: >1x10^6 frozen cells/vial, 1 mL Stability: Stable in culture over a minimum of 10 passages
| Cat. No. | CSC-RO0121 |
| Description | Ba/F3-EML4-ALK/L1196M cell line is a stably transfected cell line which expresses human EML4-ALK fusion protein with L1196M mutation in ALK. |
| Target Gene | EML4-ALK |
| Gene Species | Homo sapiens (Human) |
| Host Cell | Ba/F3 |
| Host Cell Species | Mus musculus (Mouse) |
| Stability | Stable in culture over a minimum of 10 passages |
| Application | Drug screening and biological assays |
| Growth Conditions | 37 °C, 5% CO2 |
| Quality Control | Negative for bacteria, yeast, fungi and mycoplasma. |
| Shipping | Dry ice |
| Size | >1x10^6 frozen cells/vial, 1 mL |
| Biosafety Level | 2 |
| Thawing & Subculturing Instructions | 1. Thaw cells by gently swirling in a 37°C water bath. To limit contamination, do not submerge the O-ring and cap. 2. When cells are ~70% thawed (~1 min), transfer the vial into a biosafety cabinet, and wipe the surface with 70% ethanol. Allow tube to dry completely. 3. Transfer the cells gently into a 15 mL conical tube containing 10 mL of pre-warmed culture medium (without antibiotic selection marker). Centrifuge cells at ~125 x g for 5~7 min. 4. Remove supernatant without disturbing the pellet, and resuspend cells in 1 mL culture medium (without antibiotic selection marker). Transfer cells to a 6-well plate containing ~2 mL pre-warmed growth medium (without antibiotic selection marker) or a T25 flask containing 5 mL pre-warmed culture medium (without antibiotic selection marker). 5. Incubate the culture at 37°C with 5% CO2. 6. Subculture: split saturated culture 1:4 ~ 1:6 every 3 days; seed out at about 1~3 x 10^5 cells/mL. |
| Growth Properties | Suspension, round |
| Freeze Medium | Frozen with 70% medium, 20% FBS, 10% DMSO |
| Freezing Instructions | Cells are recommended to generate additional frozen stocks at early passages. Frozen stocks should be preserved in a designated cryopreservation medium or in 70% RPMI 1640 + 20% FBS + 10% DMSO (without antibiotic selection marker). 1. Prepare the freezing medium (70% RPMI 1640 + 20% FBS + 10% DMSO, without antibiotic selection marker) fresh immediately before use. 2. Keep the freezing medium on ice and label cryovials. 3. Transfer cells to a sterile, conical centrifuge tube, and count the cells. 4. Centrifuge the cells at 250 x g for 5 minutes at room temperature and carefully aspirate off the medium. 5. Resuspend the cells at a density of at least 3 x10^6 cells/ml in chilled freezing medium. 6. Aliquot 1 ml of the cell suspension into each cryovial. 7. Freeze cells in the CoolCell freezing container overnight in a -80°C freezer. 8. Transfer vials to liquid nitrogen for long-term storage. |
| Mycoplasma | Negative |
| Format | One frozen vial containing millions of cells |
| Storage | Liquid nitrogen |
| Safety Considerations |
The following safety precautions should be observed. 1. Use pipette aids to prevent ingestion and keep aerosols down to a minimum. 2. No eating, drinking or smoking while handling the stable line. 3. Wash hands after handling the stable line and before leaving the lab. 4. Decontaminate work surface with disinfectant or 70% ethanol before and after working with stable cells. 5. All waste should be considered hazardous. 6. Dispose of all liquid waste after each experiment and treat with bleach. |
| Ship | Dry ice |
Patients diagnosed with lung cancer containing anaplastic lymphoma kinase (ALK) gene fusions experience therapeutic benefits from ALK kinase inhibitors. However, the development of acquired resistance is an inevitable challenge in treatment. Researchers utilized the EML4‐ALK Stable Cell Line to investigate proximal ALK signaling mechanisms in lung cancer cells harboring ALK gene fusions. Through affinity purification coupled with mass spectrometry and tyrosine phosphoproteomics, they delineated an ALK signaling network comprising 464 proteins. Subsequent shRNA screening identified 64 and 9 proteins whose depletion sensitized cells to crizotinib and alectinib, respectively. Knockdown of scaffolding proteins FRS2 and CC2D1A enhanced ALK inhibitor efficacy across multiple ALK fusion cell lines. These findings shed light on ALK signaling dynamics and drug resistance mechanisms, offering potential targets to enhance ALK inhibitor efficacy in lung cancer patients.
Figure 1. Researchers established an EML4‐ALK Stable Cell Line with the L1196M gate-keeper mutation, conferring resistance to crizotinib yet sensitivity to alectinib. Y27632, alone or with crizotinib, had no impact on cell viability; however, its combination with alectinib reduced IC50 across multiple cell lines, including crizotinib-resistant H3122 EML4-ALKL1196M cells. ROCK2 abundance remained consistent across tested cell lines, irrespective of crizotinib sensitivity. Additionally, HSP90 and CDC37 inhibition reduced H3122 cell viability. (Zhang G, et al., 2016)
A: HEK293 cells were likely selected for their high transfection efficiency and suitability for stable transgene expression, facilitating the study of LAG3 biology, immune regulation, and therapeutic antibody development.
A: Stability was likely confirmed through methods such as flow cytometry, immunoblotting, or functional assays measuring LAG3-mediated signaling, with continuous selection pressure applied.
A: Characterization may involve analysis of LAG3 membrane localization, interaction with binding partners such as MHC class II molecules or CD4, downstream signaling pathways, and functional implications in T cell activation, exhaustion, or immune checkpoint blockade.
A: Quality control likely included confirmation of LAG3 expression levels, validation of its functional activity, assessment of off-target effects, and validation of phenotypic changes associated with LAG3 modulation.
A: Comparative analysis with primary immune cells or in vivo models helps validate the relevance of LAG3 expression in immune checkpoints, tumor immune evasion, and its potential as a target for cancer immunotherapy or autoimmune disease treatment.
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Unmatched consistency! The Human EML4-ALK/L1196M Stable Cell Line in Ba/F3 cells delivers reliable EML4-ALK/L1196M fusion protein expression, ensuring robust results in my cancer research.
Empowering advanced studies! With stable expression of the oncogenic EML4-ALK/L1196M fusion, I can explore ALK inhibitor resistance mechanisms and novel targeted therapies with confidence, driving scientific breakthroughs.
Exceptional performance! This cell line surpasses expectations, providing a solid foundation for investigating EML4-ALK/L1196M-targeted therapies and precision medicine approaches in ALK-positive cancers.
Streamlining research processes! Its stable expression simplifies experimental workflows, enhancing efficiency and enabling in-depth data analysis with ease.
An indispensable asset! The Human EML4-ALK/L1196M Stable Cell Line has revolutionized my research, offering valuable insights into EML4-ALK/L1196M-driven oncogenesis and potential therapeutic interventions for ALK-positive tumors.
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