Fig. 1

Mig-6 is overexpressed in gefitinib-resistant PC9/GR cell lines compared with parental PC9 cell lines. a Mig-6 expression was examined by immunofluorescence staining of endogenous Mig-6 (red) in parental PC9 cells and in PC9/GR cells (Original magnification, × 100 as displayed in the figures), lower panel is optical microscopy images of PC9 and PC9/GR cells (scale bar represents 100 μm). b Western blot experiments for proteins related to epidermal growth factor receptor (EGFR) signaling and the epithelial-mesenchymal transition (EMT) in PC9 and PC9/GR cells. The expressions Mig6, p-EGFR (Tyr1068), p-EGFR (Tyr1045), EGFR, c-MET, p-AKT, AKT, p-ERK, ERK, E-cadherin, Zo-1 and Vimentin were evaluated. β-actin was used as a loading control. c Phos-tag immunoblot analysis of Mig-6 with or without phosphatase. To check the phosphorylated status of Mig-6, the lysates we employed phos-tag SDS-PAGE. The lysate was loaded in 6% Acrylamide 100uM Mn²⁺-Phos-tag™ Acrylamide. To confirm that those slower migrating bands are really phosphorylated species of Mig-6, we treated a part of the lysate with protein phosphatase for 9 h. d RT-PCR analysis. mRNA levels of EGFR, E-cadherin, Vimentin, and c-MET was evaluated in PC9 and PC9/GR. GAPDH was used as an internal control. Uncropped blots were shown in Additional file 1: Fig. S1