KC-3905

Raji-EGFR-High-Cell-Line

35006

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Background of Raji-EGFR-High-Cell-Line

EGFR is a transmembrane glycoprotein that is one of four members of the ErbB family of tyrosine kinase receptors. Activation of EGFR leads to autophosphorylation of the receptor tyrosine kinase, which initiates downstream signaling cascades involved in the regulation of cell proliferation, differentiation, and survival. The abnormal activation of EGFR through a variety of mechanisms, such as receptor overexpression, mutation, ligand-dependent receptor dimerization, ligand-independent activation, is associated with the occurrence and development of a variety of human tumors. Inhibition of EGFR is one of the key targets of cancer chemotherapy.

Specifications

Catalog Number	KC-3905
Cell Line Name	Raji-EGFR-High-Cell-Line
Host Cell Line	Raji
Description	Stable Raji cell line expressing exogenous human EGFR 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 + 400 µg/mL Hygromycin B
Selection Marker	Hygromycin B
Morphology	Mostly single, round (some polymorph) cells in suspension
Subculture	Split saturated culture 1:3 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 24-36 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

Raji human EGFR High cell line was generated using a lentiviral vector expressing the human EGFR sequence.

Characterization

Figure1: Characterization of EGFR overexpression in Raji stable clones using FACS.

Figure2: Characterization of EGFR and its mutants overexpressing in Raji stable clones using PCR sequencing

Cell Resuscitation

Prewarm culture medium (RPMI1640 + 10% FBS + 400 µg/mL Hygromycin B)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 1:2 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% RPMI-1640 + 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

ingh D, Attri BK, Gill RK, Bariwal J. Review on EGFR Inhibitors: Critical Updates. Mini Rev Med Chem. 2016;16(14):1134-66. doi: 10.2174/1389557516666160321114917. PMID: 26996617.
Yarden Y, Shilo BZ. SnapShot: EGFR signaling pathway. Cell. 2007 Nov 30;131(5):1018. doi: 10.1016/j.cell.2007.11.013. PMID: 18045542.