Epidermal growth factor receptor (EGFR) is a type I transmembrane glycoprotein and member of the receptor protein kinase superfamily. EGFR, along with its ligands, play critical roles in a wide variety of cellular functions, including cell proliferation, differentiation, motility and survival. Ligands for EGFR include EGF, TGF-?, amphiregulin, betacellulin, heparin-binding EGF-like growth factor, GP30 and vaccinia virus growth factor (1). The full length human EGFR contains 1186 amino acid (aa) residue with a 621 aa extracellular domain, a 23 aa transmembrane domain, and a 542 aa cytoplasmic domain (2). Ligand binding induces EGFR homo and hetero-dimerization, resulting in kinase activation, tyrosine phosphorylation and cell signaling (3). EGFR signaling is known to induce the MAPK, Akt, and JNK signaling pathways (4). EGFR is expressed by many types of epithelial and endothelial cells and frequently upregulated in many types of cancers (5).
THAWING AND CULTURING
• Remove the cell vial from liquid nitrogen tank and thaw cells quickly in a 37°C water bath
• Transfer the cells to a 15 ml centrifuge tube and slowly add 5 ml of pre-warmed complete growth medium
• Centrifuge the cells at 200x g for 5 min
• Remove the supernatant
• Resuspend cell pellet with 7 ml of complete growth medium and transfer cells to a T25 flask
• Incubate cells in an incubator with 5% CO2 at 37°C
• Split the cells twice a week or as needed
Detection of human EGFR expression on human EGFR-CT26 stable cells using monoclonal antibodies specific for human EGFR.
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