KC-6067

Raji-EGFR-PDL1 Cell Line

63942

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Background of Raji-EGFR-PDL1 Cell Line

EGFR (Epidermal Growth Factor Receptor) is a Protein Coding gene. Diseases associated with EGFR include Neonatal Nephrocutaneous Inflammatory Syndrome and Lung Cancer. Among its related pathways are Apoptotic Pathways in Synovial Fibroblasts and Signaling by EGFR in Cancer.MET (MET Proto-Oncogene, Receptor Tyrosine Kinase) is a Protein Coding gene. Diseases associated with MET include Renal Cell Carcinoma, Papillary, 1 and Arthrogryposis, Distal, Type 11. Among its related pathways are Apoptotic Pathways in Synovial Fibroblasts and GPCR Pathway. Gene Ontology (GO) annotations related to this gene include transferase activity, transferring phosphorus-containing groups and protein tyrosine kinase activity.
PD-L1, also called human programmed cell death ligand 1, is a transmembrane protein that plays a major role in suppressing the immune system during particular events such as pregnancy, tissue allografts, autoimmune disease, virus infection, and cancer. PD-L1 binds to its receptor PD-1 on activated T cells, B cells, and myeloid cells, to modulate activation or inhibition. Upregulation of PD-L1 can allow the cancer cell to evade the host immune system.

Specifications

Catalog Number	KC-6067
Cell Line Name	Raji-EGFR-PDL1 Cell Line
NCBI/UniProt Accession Number	NM_005228, Q9NZQ7
Clone Number	1#
Host Cell Line	Raji
Description	Stable Raji cell line expressing exogenous EGFR and PDL1 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% basal medium+20% FBS+10% DMSO
Propagation Medium	RPMI 1640+10% FBS + 5μg/mL BSD + 400μg/mL Hygromycin B
Selection Marker	Hygromycin B, BSD
Morphology	Epithelial-like
Subculture	Split saturated culture 1:4-1:6 every 2-3 days
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

Raji-EGFR-PDL1 cell line was generated using a lentiviral vector expressing the EGFR-PDL1 sequence.

Characterization

Figure 1: Characterization of EGFR-PDL1 overexpression in the Raji-EGFR-PDL1 stable clone using FACS.

Figure 2: Characterization of EGFR-PDL1 in the Raji-EGFR-PDL1 stable clone using PCR sequencing.

Cell Resuscitation

Pre-warm complete culture medium (basal medium and 10% FBS) in a 37°C water bath.
Rapidly thaw the cryovial in a 37°C water bath for 1-2 minutes with gentle agitation.
Transfer the vial to a biosafety cabinet, and disinfect the exterior with 70% ethanol.
Aseptically transfer the cell suspension dropwise into a sterile centrifuge tube containing 9.0 mL of pre-warmed complete medium.
Centrifuge at approximately 125 × g for 5–7 minutes at room temperature, carefully aspirate the supernatant without disturbing the cell pellet.
Gently resuspend the pellet in an appropriate volume of complete medium and transfer the suspension into a T25 flask.
Incubate the flask in a 37°C in a humidified 5% CO₂ incubator.
Assess cell viability and morphology after 24 hours. If cells appear healthy, replace the medium with fresh medium supplemented with the appropriate selective antibiotic.
Subculture the cells at a ratio of 1:4-1:6 every 2-3 days upon reaching 80%–90% confluency.

Cell Freezing

Prepare the freezing medium (70% basal medium, 20% FBS and 10% DMSO) freshly before use.
Pre-chill the freezing medium on ice and label the cryovials accordingly.
Transfer the cell suspension to a sterile conical tube and perform a cell count to determine total viability and density.
Centrifuge the cells at 250×g for 5 minutes at room temperature; carefully aspirate the supernatant.
Gently resuspend the cell pellet in chilled freezing medium, ensuring a minimum cell density of 3×10⁶ cells/mL.
Aliquot 1 mL of the cell suspension into each pre-labeled cryovial.
Place the cryovials into a CoolCell® container and store at -80°C overnight for controlled-rate cooling.
Transfer the cryovials to the liquid nitrogen for long-term storage the following day.

References

1.Sunshine, J. & Taube, J. M. PD-1/PD-L1 inhibitors. Current Opinion in Pharmacology 23, 32ÿ38 (2015). 2. Boussiotis, V. A. Molecular and Biochemical Aspects of the PD-1 Checkpoint Pathway. N Engl J Med 375, 1767ÿ 1778 (2016). 3. Topalian, S. L. et al. Safety, activity, and immune correlates of anti-PD-1 antibody in cancer. N Engl J Med 366, 2443ÿ2454 (2012).
2.Campbell P, Morton PE, Takeichi T, Salam A, Roberts N, Proudfoot LE, Mellerio JE, Aminu K, Wellington C, Patil SN, Akiyama M, Liu L, McMillan JR, Aristodemou S, Ishida-Yamamoto A, Abdul-Wahab A, Petrof G, Fong K, Harnchoowong S, Stone KL, Harper JI, Irwin McLean WH, Simpson MA, Parsons M, McGrath JA. Epithelial inflammation resulting from an inherited loss-of-function mutation in EGFR. J Invest Dermatol. 2014 Oct;134(10):2570-2578.

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.