Fuji super rx n x ray film

Cellular lysates were prepared using the methods outlined in previous studies [18 , 19 (link)]. Proteins were resolved on sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis gel and transferred to Immobilon PVDF membranes. The blots were blocked with 5% BSA for 1 h at room temperature and then probed with rabbit antihuman antibodies against EGFR, PI3K, Akt, IKKα/β, IκB, and p65 (1 : 1000) for 1 h at room temperature. After 3 washes, the blots were subsequently incubated with donkey anti-rabbit peroxidase-conjugated secondary antibody (1 : 3000) for 1 h at room temperature. The blots were visualized using enhanced chemiluminescence and FUJI Super RX-N X-RAY film (Fujifilm Corporation, Tokyo, Japan). Quantitative data were obtained using TotalLab 2.0 software (Nonlinear Dynamics, BioSystematica, UK).

Wound-healing migration assay and western blot analysis were performed as previously described with following modifications [46 (link), 82 (link)]. After transfer, the blots were blocked with 5% BSA at room temperature for 1 hr. and then probed with different rabbit anti-human antibodies that are against PI3K, Akt, IKKα/β, IκB, p65, EGFR, ICAM-1 (1:1000) at room temperature for 1 hr. The blots were visualized by enhanced chemiluminescence which can be detected by FUJI Super RX-N X-RAY film (Fujifilm Corporation, Tokyo, Japan). Quantitative data were obtained using a computing densitometer and Image Quant software (Molecular Dynamics, Sunnyvale, CA, USA).