Cat.No. : CSC-RO0418 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-RO0418 |
| Description | This cell line is engineered to stably overexpress exogenous human IGF1R protein. |
| Target Gene | IGF1R |
| 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 |
| Storage | Liquid nitrogen |
| 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 |
Triple-negative breast cancer (TNBC) is a highly metastatic disease that currently lacks effective preventive and therapeutic strategies. The insulin-like growth factor 1 receptor (IGF1R) and focal adhesion kinase (FAK) signaling pathways play a role in many developmental processes, and alterations in both are associated with many common pathological diseases. Overexpression of IGF1R and FAK is closely associated with metastatic breast tumors. The studies here show that IGF1R overexpression promotes the migratory and invasive behavior of TNBC cell lines. Most importantly, IGF1R-driven migration and invasion are mainly mediated by FAK activation and can be inhibited using pharmacological inhibitors of FAK. These results may improve the current understanding of the basic molecular mechanisms of TNBC metastasis and provide a strong rationale for simultaneously targeting IGF1R and FAK as a therapeutic approach for mesenchymal TNBC.
The researchers used stably transfected EV control (Hs578T-EV) or IGF1R overexpressing Hs578T cells (Hs578T-IGF1R(+/+)) to test the potential requirement of FAK in IGF1R-mediated migration and invasion. Immunoblot analysis showed that IGF1R overexpression increased FAK activity and ZEB-1 expression, accompanied by a decrease in epithelial biomarkers E-cadherin and ZO-1 (Figure 1A). In addition, quantitative real-time PCR experiments showed that IGF1R overexpressing cells expressed lower levels of E-cadherin mRNA and higher levels of vimentin mRNA compared with IGF1R-deficient cells (Figure 1B). Consistent with the absence of active FAK protein, Hs578T-EV cells were unresponsive to both PF228 and PF878 treatment; however, IGF1R-overexpressing Hs578T cells exhibited reduced clonogenicity after treatment with both inhibitors (Figure 1C). Similarly, FAK inhibitor treatment did not affect the migration of IGF1R-deficient Hs578T cells, but IGF1R-overexpressing Hs578T cells showed increased cell migration that was inhibited in the presence of both FAK inhibitors (Figure 1D). Unlike IGF1R-deficient cells that continued to invade in the presence of PF228 and PF878 inhibitors, IGF1R overexpression induced increased invasiveness in TNBC cells, which was also effectively reduced by FAK inhibitors (Figure 1E). Inhibitor treatment had no significant effect on cell proliferation in either cell line (Figure 1F). Taken together, these results suggest that FAK is an important molecule mediating clonogenicity as well as the pro-migratory and pro-invasive effects of IGF1R in TNBC cells.
Figure 1. Inhibition of FAK abrogates IGF1R-mediated colony formation, migration, and invasion in TNBC cells. (Taliaferro-Smith L T, et al. 2015)
Creative Biogene has successfully constructed IGF1R over-expressing stable cell lines with Hs578T, CHO-K1, HEK293, BaF3, TE-1, CHO, HeLa, 1321N1, U2OS, etc. These IGF1R stable cell lines have excellent in vitro assay sensitivity and reproducibility. Therefore, conducting experiments with our IGF1R stable cell lines saves time and resources while providing more accurate results.
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The expression levels of IGF1R in these BaF3 cells are very high, which makes our signaling pathway studies much simpler. The robust expression ensures that our assays are highly sensitive, which improves the accuracy of our data.
We have found the Human IGF1R Stable Cell Line - BaF3 to be very easy to culture and maintain. The cells grow vigorously and adapt well to a variety of conditions, which reduces the time we need to spend on maintenance.
In comparison to other IGF1R cell lines we've used, the Human IGF1R Stable Cell Line - BaF3 offers an excellent cost-to-benefit ratio. The cells' high performance and low maintenance requirements translate into cost savings for our lab.
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