Skip to main content
. 2021 Sep 10;11:664236. doi: 10.3389/fonc.2021.664236

Figure 1.

Figure 1

Construction and expression of 806 CAR and EGFR mutant cell lines. (A) Schematic diagram of vector map of 806 CAR containing the 4-1BB co-stimulatory domain. (B) CAR surface expression in primary human CD4+ and CD8+ T cells. Human T cells were simulated for 24 h with anti-CD3/anti-CD28 T-cell activating beads and transduced with CAR transgenes, and CAR expression was analyzed by flow cytometry using biotinylated goat-anti-mouse (806, C225, and CD19 CARs) and goat-anti human F(ab)2 fragment-specific antibodies (2173 CARs) followed by secondary staining with streptavidin-APC. (C) Schematic showing targeted missense mutations in the extracellular domain of EGFR, EGFRR108K/G, EGFRA289D/T/V, EGFRG598V, and splice variant EGFRvIII. (D) Schematic of lentiviral vector co-expressing CFP and wtEGFR or EGFR mutant. (E) Flow-based analysis of endogenous and ectopically expressed EGFR in U87MG, GSC5077, and K562 cell lines using the cetuximab antibody. (F) U87MG, U87MG-EGFR, and GSC5077-EGFR expression of EGFRvIII. (G) U87MG-EGFR, GSC5077-EGFR, and K562 cell lines were transduced with a lentiviral vector co-expressing CFP and indicated EGFR missense mutations and sorted by CFP expression using fluorescent-activated cell sorting.