(A) The chart represents the number of viable cells in PC9, PC14, and AXL-positive cells sorted from PC9 upon treatment with indicated doses of Erlotinib. Values are normalized relative to vehicle-treated cell (control). Cells were grown for 120 hr in the presence of the drug; the number of cells was estimated upon staining with the crystal violet, de-staining in 100 µl of 10% acetic acid and reading absorbance at 590 nm. Diamonds and black bars represent single-point measurements and the mean, respectively (n = 6); (*p< 0.05, **<0.005, ***<0.0005, unpaired t-test). (B) AXL-positive cells are characterized by mesenchymal features such as an increase in stress fibers. AXL-negative and AXL-positive cells sorted from PC9 were stained F-actin with Phalloidin (green). DAPI (blue) was used as a counter-stain. (C) The charts represent the relative expression of the indicated mesenchymal signature genes in H1650, H1650-M3, and cells sorted based on surface expression of AXL from H1650 and PC9. Expression of an indicated mRNA in the AXL-positive cells was calculated relative to its expression in AXL-negative control cells mRNA expression was quantified by SYBR-green-based RT-qPCR. Each bar represents mean ± SD of three replicates from two independent experiments (*p< 0.05, **<0.005, ***<0.0005, ****<0.00005, unpaired t-test). (D) Expression of mesenchymal signature genes VIM, TWIST, and ZEB1 in FACS-sorted AXL-negative (blue) and AXL-positive (red) cells from five human primary NSCLC tumors. mRNA expression was quantified by Cells to CT one-step SYBR-green-based RT-qPCR. The expression of an indicated mRNA in the AXL-positive cells was calculated relative to its expression in AXL-negative cells from the respective tumor. Each dot represents mean ± SD of three replicates.