KC-5098

CHOK1-cyno-FGFR2b Cell Line

54528

Home » CHOK1-cyno-FGFR2b Cell Line

Background of CHOK1-cyno-FGFR2b Cell Line

The family of fibroblast growth factor receptors (FGFRs) includes four transmembrane receptor tyrosine kinases (FGFR1–4) involved in the regulation of crucial biological processes, such as proliferation, migration, survival, and differentiation. The FGFR proteins are involved in a wide array of pathways known to play a signficant role in cancer, it can stimulate the transformation and proliferation of tumor cells and angiogenesis. Fibroblast growth factor receptor 2 (FGFR2) fusion proteins (FFs) are oncogenic drivers in 10-15% of intrahepatic cholangiocarcinoma (iCCA). Bemarituzumab, an FGFR2b inhibitor, is the first-in-class humanized, non-fructogenated immunoglobulin G1 monoclonal antibody. It acts on FGFR2b overexpressing tumors by inhibiting FGFR2b signaling and enhancing antibody-dependent cell-mediated cytotoxicity (ADCC).

Specifications

Catalog Number	KC-5098
Cell Line Name	CHOK1-cyno-FGFR2b Cell Line
Clone Number	3-8#
Host Cell Line	CHOK1
Description	Stable CHOK1 cell line expressing exogenous cyno FGFR2b 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 + 10μg/ml Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 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 24 hours
Mycoplasma Status	Negative

Cell Line Generation

CHOK1-cyno-FGFR2b Cell Line was generated using a lentiviral vector expressing the cyno FGFR2b sequence.

Characterization

Figure 1: Characterization of cyno FGFR2b overexpression in the CHOK1 cyno FGFR2b stable clone using FACS.

Figure 2: Characterization of cyno FGFR2b and its mutants overexpressing in CHOK1 stable clones using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 10μg/mL Puromycin)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 1:4-1:8 every 2-3 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. Xiang H, Chan AG, Ahene A, Bellovin DI, Deng R, Hsu AW, Jeffry U, Palencia S, Powers J, Zanghi J, Collins H. Preclinical characterization of bemarituzumab, an anti-FGFR2b antibody for the treatment of cancer. MAbs. 2021 Jan-Dec;13(1):1981202. doi: 10.1080/19420862.2021.1981202. PMID: 34719330; PMCID: PMC8565817.
2. Rosato B, Ranieri D, Nanni M, Torrisi MR, Belleudi F. Role of FGFR2b expression and signaling in keratinocyte differentiation: sequential involvement of PKCδ and PKCα. Cell Death Dis. 2018 May 1;9(5):565. doi: 10.1038/s41419-018-0509-x. PMID: 29752438; PMCID: PMC5948219.
3. Ahmadvand N, Lingampally A, Khosravi F, Vazquez-Armendariz AI, Rivetti S, Jones MR, Wilhelm J, Herold S, Barreto G, Koepke J, Samakovlis C, Carraro G, Zhang JS, Al Alam D, Bellusci S. Fgfr2b signaling is essential for the maintenance of the alveolar epithelial type 2 lineage during lung homeostasis in mice. Cell Mol Life Sci. 2022 May 19;79(6):302. doi: 10.1007/s00018-022-04327-w. PMID: 35587837; PMCID: PMC9120111.