KC-4097

Ba/F3-FGFR3IIIc Cell Line

41803

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Background of Ba/F3-FGFR3IIIc Cell Line

This gene encodes a member of the fibroblast growth factor receptor (FGFR) family, with its amino acid sequence being highly conserved between members and among divergent species. FGFR family members differ from one another in their ligand affinities and tissue distribution. A full-length representative protein would consist of an extracellular region, composed of three immunoglobulin-like domains, a single hydrophobic membrane-spanning segment and a cytoplasmic tyrosine kinase domain. The extracellular portion of the protein interacts with fibroblast growth factors, setting in motion a cascade of downstream signals, ultimately influencing mitogenesis and differentiation. This particular family member binds acidic and basic fibroblast growth hormone and plays a role in bone development and maintenance. Mutations in this gene lead to craniosynostosis and multiple types of skeletal dysplasia.
Ba/F3 cell, a murine interleukin-3 dependent pro-B cell line, is a popular system for exploring both kinases and their inhibitors, because some protein kinases can render the Ba/F3 cells to be depended on the activation of the kinases instead of IL-3 supplement, while their inhibitors can antagonize the kinase-dependent growth effects.

Specifications

Catalog Number	KC-4097
Cell Line Name	Ba/F3-FGFR3IIIc Cell Line
NCBI/UniProt Accession Number	NM_000142.5
Clone Number	2-10#
Host Cell Line	Ba/F3
Description	Stable Ba/F3 clone expressing exogenous FGFR3IIIc gene.
Quantity	Two vials of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10%FBS+1µg/mL Puromycin+8ng/mL mouse IL-3
Selection Marker	Puromycin
Morphology	Mostly single, round (some polymorph) cells in suspension
Subculture	Split saturated culture 1:10 every 3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

Ba/F3 FGFR3IIIc cell Line was generated using retrovirus vector expressing FGFR3IIIc sequence.

Characterization

Figure1: Characterization of FGFR3IIIc overexpressing in Ba/F3 stable clone using FACS.

Figure 2: Characterization of FGFR3IIIc overexpressing in Ba/F3-FGFR3IIIc stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640+10%FBS+1µg/mL Puromycin+8ng/mL mouse IL-3)in a 37°C water bath.
2. Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
3. Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
4. Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
5. Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
6. Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
7. Incubate the flask at 37°C, 5% CO₂ incubator.
8. Split saturated culture 1:10 every 3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% RPMI1640 + 20% FBS + 10% DMSO) 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×g for 5 minutes at room temperature and carefully aspirate off the medium.
5. Resuspend the cells at a density of at least 3×10⁶ 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.

References

1. Yang Y, Li DZ. FGFR3 gene mutations in Chinese cases of thanatophoric dysplasia type 1. Fetal Diagn Ther. 2009;26(2):90-2. doi: 10.1159/000238120. Epub 2009 Sep 11. PMID: 19752524.
2.Quintero-Rivera F, El-Sabbagh Badr R, Rao PN. FGFR3 amplification in the absence of IGH@-FGFR3 fusion t(4;14) in myeloma. Cancer Genet Cytogenet. 2009 Nov;195(1):92-3. doi: 10.1016/j.cancergencyto.2009.06.018. PMID: 19837276.