Equivalent amounts of protein (50 μg, determined by Bradford assays) were resolved on 10% SDS-polyacrylamide gels. The protein was electrotransferred to a polyvinylidene difluoride membrane (Millipore, Bedford, MA), which was then blocked in a solution of 5% (wt/vol) skim milk powder in PBS (pH 7.5) for 1 hour at room temperature and probed overnight at 4°C with anti-integrin α5 (H-104, 1:300; Santa Cruz Biotechnology, Fremont, CA), anti-integrin β1 (EP1041Y, 1:200; Abcam, Cambridge, MA) or anti-β-actin (1:1000; Cell Signaling Technology, Danvers, MA) antibodies. After membranes were washed with PBS, horseradish peroxidase (HRP)–conjugated goat anti-rabbit secondary antibodies (1:2000; Cell Signaling Technology) were incubated with the membranes for 1 hour at room temperature. Finally, proteins were detected by enhanced chemiluminescence (GE Healthcare, Piscataway, NJ) and visualized after exposure to autoradiographic film (Kodak, Rochester, NY). Protein levels were quantified by densitometry and normalized to β-actin levels.