Whole-cell lysates were prepared for SDS-PAGE as described previously (11 (link)). Briefly, 10–20 μg of protein were separated by SDS-PAGE, transferred to 0.45 μm polyvinylidene difluoride membranes (Bio-Rad), blocked with 5% nonfat dry milk in 1× TBS/0.1% Tween-20, and then probed with ezrin (Cell Signaling Technology, catalog no. 3145), radixin (Cell Signaling Technology, catalog no. C4G7), moesin (Cell Signaling Technology, clone Q480 catalog no. 3150), phospho-ezrin-radixin-moesin (Cell Signaling Technology, catalog no. 3726), phospho-Serine473 Akt (Cell Signaling Technology, catalog no. 9271), total Akt (Cell Signaling Technology, catalog no. 9272), phospho-Serine136 Bad (D25H8, Cell Signaling Technology, catalog no. 4366), total Bad (Cell Signaling Technology, catalog no. 9292), phospho-Serine536 p65 NFκB (Cell Signaling Technology, catalog no. 3031), total p65 NFκB (Cell Signaling Technology, catalog no. 8242), survivin (Cell Signaling Technology) or γ-tubulin (Sigma-Millipore, catalog no. T5326), with the appropriate secondary antibodies (Cell Signaling Technology).
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