KC-5753

Ba/F3-EGFR-K728R Cell Line

58844

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Background of Ba/F3-EGFR-K728R Cell Line

EGFR (Epidermal Growth Factor Receptor) is a receptor tyrosine kinase that is a critical driver of cell proliferation and survival. Its aberrant activation, through overexpression or mutations, is a well-established oncogenic event in multiple cancers, particularly non-small cell lung cancer (NSCLC). This has made EGFR a premier drug target, leading to the development of tyrosine kinase inhibitors (TKIs). A central focus of current EGFR research is understanding and overcoming the inevitable drug resistance that arises to these targeted therapies.

Specifications

Catalog Number	KC-5753
Cell Line Name	Ba/F3-EGFR-K728R Cell Line
Clone Number	NA
Host Cell Line	Ba/F3
Description	Stable Ba/F3 cell line expressing exogenous EGFR-K728R gene
Quantity	One vial 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% RMPI1640 + 20% FBS + 10% DMSO
Propagation Medium	RPMI1640+10%FBS
Selection Marker	NA
Morphology	Mostly single, round (some polymorph) cells in suspension
Subculture	Split saturated culture 1:4-1:10 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 20 hours
Mycoplasma Status	Negative

Cell Line Generation

Ba/F3-EGFR-K728R Cell Line was generated using a retrovirus vector expressing the EGFR-K728R sequence.

Characterization

Figure 1: Characterization of EGFR-K728R overexpression in the Ba/F3-EGFR-K728R stable clone using PCR.

Cell Resuscitation

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

Cell Freezing

Prepare the freezing medium (70% RPMI1640 + 20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage.

References

Arrieta O, Cardona AF, Corrales L, Campos-Parra AD, Sánchez-Reyes R, Amieva-Rivera E, Rodríguez J, Vargas C, Carranza H, Otero J, Karachaliou N, Astudillo H, Rosell R; CLICaP. The impact of common and rare EGFR mutations in response to EGFR tyrosine kinase inhibitors and platinum-based chemotherapy in patients with non-small cell lung cancer. Lung Cancer. 2015 Feb;87(2):169-75. doi: 10.1016/j.lungcan.2014.12.009. Epub 2014 Dec 18. PMID: 25558790.
Wheeler SE, Egloff AM, Wang L, James CD, Hammerman PS, Grandis JR. Challenges in EGFRvIII detection in head and neck squamous cell carcinoma. PLoS One. 2015 Feb 6;10(2):e0117781. doi: 10.1371/journal.pone.0117781. PMID: 25658924; PMCID: PMC4320077.

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.