Fig. 5. Expression and localization of uPAR and ß1-integrin to caveolae are downregulated in AS-cav-1 HCT 116 cells. (A) Control and AS-cav-1 HCT 116 cells were solubilized in lysis buffer containing 1% Triton X-100, 60 mM octylglucoside. 30 µg protein were analysed by SDS-PAGE and immunoblotted with anti-human-uPAR antibodies. Immunoblots are shown in triplicate and are representative of at least three experiments. (B) Equal amounts of protein for control and AS-cav-1 HCT 116 cells were subjected to subcellular fractionation on a sucrose gradient after homogenization in sodium-carbonate buffer, pH 11.0. Fractions were collected from the top of the gradient and equal-volume aliquots of fractions 2-11 were analysed by SDS-PAGE and immunoblotted with anti-human-uPAR antibody. (C) Control and AS-cav-1 HCT 116 cells were solubilized in lysis buffer containing 1% Triton X-100, 60 mM octylglucoside. 20 µg protein were analysed by SDS-PAGE and immunoblotted with anti-human-ß1-integrin antibody. (D) Equal amounts of protein for control and AS-cav-1 HCT 116 cells were subjected to subcellular fractionation on a sucrose gradient after homogenization in sodium-carbonate buffer, pH 11.0. Fractions were collected from the top of the gradient and equal-volume aliquots of fractions 2-11 were analysed by SDS-PAGE and immunoblotted with anti-human-ß1-integrin antibodies. Immunoblots are representative of at least three experiments.