Cat.No. : CSC-RO1109M 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-RO1109M |
| Description | Ba/F3-KIT cell line is engineered to stably overexpress human KIT. |
| Target Gene | KIT |
| Gene Species | Homo sapiens (Human) |
| Alias | C-Kit, CD117, MASTC, PBT, SCFR |
| 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 |
The maintenance of adult long-term hematopoiesis relies on the preservation of hematopoietic stem/progenitor cells (HSPC) within specialized niches in the bone marrow (BM). These niches tightly regulate the equilibrium between quiescence, self-renewal, and differentiation of HSPC. Researchers utilized the Human KIT Stable Cell Line-Ba/F3 to investigate the intrinsic responsiveness of HSPC to niche factors. By expressing membrane-targeted C3G (C3G-F), a Rap1 GTP/GDP exchanger, they observed enhanced proliferation and differentiation of HSPC in response to stem cell factor (SCF) and thrombopoietin (TPO). This prolonged activation of c-Kit receptor and downstream signaling through SCF ligation sheds light on the crucial role of basal Rap1 activation in maintaining HSPC in bone marrow niches, crucial for long-term adult hematopoiesis.
Figure 1. Increased basal Rap1GTP level in Human KIT Stable Cell Line-Ba/F3 leads to prolonged stem cell factor (SCF)‐mediated c‐Kit receptor activation and downstream signaling. Protein expression and cell surface c‐Kit were examined, revealing altered levels. Effects of SCF stimulation on cell viability and migration were assessed. Signaling dynamics were evaluated through immunoprecipitation and immunoblotting. (Imai T, et al., 2019)
A: Ba/F3 cells were chosen for establishing the stable cell line expressing human KIT due to their lack of endogenous KIT expression and their responsiveness to KIT activation. Ba/F3 cells offer a suitable background for studying KIT signaling and drug sensitivity.
A: Verification and maintenance of stability and expression level of human KIT in the Ba/F3 stable cell line involved stable transfection, clonal selection, and validation of expression using techniques such as qPCR and western blot analysis. Continuous culture under appropriate conditions ensured stable expression.
A: Functional characterization of human KIT in the Ba/F3 stable cell line focused on its kinase activity and response to KIT inhibitors. This included assessing the phosphorylation of downstream signaling molecules such as STAT5 and ERK, as well as evaluating sensitivity to KIT inhibitors like imatinib using cell viability assays.
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An invaluable resource! The Human KIT Stable Cell Line has significantly enhanced my research capabilities, offering valuable insights into KIT-mediated signaling networks and potential therapeutic strategies for KIT-associated diseases.
Impressive reliability. This cell line surpasses expectations, serving as a robust platform for studying KIT-targeted therapies and potential interventions for KIT-driven malignancies. Its stable expression simplifies experimental workflows, facilitating efficient data collection and analysis, and accelerating discoveries in hematology and cancer research.
With stable KIT expression, I can investigate signaling pathways downstream of KIT and KIT-mediated cellular processes with confidence, advancing our understanding of hematopoietic disorders and oncogenesis. The Human KIT Stable Cell Line in Ba/F3 cells ensures consistent expression of KIT, providing reliable results in my research on hematopoiesis and cancer.
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