Cat.No. : CSC-RO0129 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-RO0129 |
| Description | Ba/F3-EGFR-DEL19 cell line is a stably transfected cell line which expresses human epidermal growth factor receptor (EGFR) with DEL19 mutation. |
| Target Gene | EGFR |
| 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 |
Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) are effective drugs for the treatment of non-small cell lung cancer (NSCLC) harboring EGFR-activating mutations. Both the second-generation EGFR-TKI afatinib and the third-generation EGFR-TKI osimertinib are standard therapies for patients with this type of cancer. Each drug has a different binding site for the tyrosine kinase domain of EGFR. This study examined the efficacy of single and combined TKI therapy against Ba/F3 cells harboring the EGFR-activating Del19 mutation using an in vitro growth inhibition assay. Treatment with afatinib or osimertinib alone significantly inhibited cell proliferation of Ba/F3 cells, but direct sequencing determined that resistant cells eventually emerged with secondary EGFR mutations (T790M or C797S, respectively). Notably, the combination of afatinib and osimertinib eradicated Ba/F3 cells without the development of resistance.
Figure 1. Afatinib and osimertinib combination therapy eradicates Ba/F3 cells with the EGFR-activating mutation Del19. (A) EGFR-DEL19 Ba/F3 cells were treated with the indicated concentrations of afatinib or osimertinib, and cell viability was measured after 3 days. (B) Overview of the experimental approach used to establish drug-resistant clones. (C) The number of drug-resistant clones was calculated per 50 wells for each treatment. (D) EGFR sequence chromatograms of EGFR-DEL19 Ba/F3 cells and afatinib-resistant Ba/F3 AR2 cells. (E) EGFR sequence chromatograms of EGFR-DEL19 Ba/F3 cells and osimertinib-resistant Ba/F3 OR5 cells. (Yonesaka K, et al., 2019)
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We are impressed by the high transfection efficiency of this cell line. It significantly increases our experimental throughput and allows for more detailed analysis of the EGFR signaling pathway with minimal optimization.
This stable cell line has proven to be extremely valuable for our drug screening assays. The EGFR-DEL19 mutation is clinically relevant, and having such a robust, reproducible model improves our ability to identify potential therapeutic compounds.
From thawing to routine maintenance, working with the Human EGFR-DEL19 Stable Cell Line-Ba/F3 has been straightforward and hassle-free. The clear instructions and superior cell viability post-thaw make it user-friendly, even for newcomers in the lab.
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