Integrin α₂β₁ binding to biotinylated decorin immobilized on streptavidin-Sepharose.

Glycoproteins were isolated from lysed platelet membranes by affinity chromatography on concanavalin A-Sepharose. Biotinylated decorin was added to an aliquot of platelet glycoproteins and incubated for 2 hours at room temperature. Then, streptavidin-Sepharose was added to the sample and incubated for a further hour. As a negative control, an aliquot of membrane glycoproteins was incubated with buffer instead of biotinylated decorin, followed by addition of streptavidin-Sepharose. Proteins recovered from the control sample (buffer) and from the sample incubated with biotinylated decorin (DCN), together with an aliquot of the membrane glycoproteins preparation (MEMB), were separated by SDS-PAGE on a 7.5% acrylamide gel and transferred to nitrocellulose. Filters were probed with an antibody against the α2 subunit of integrin α₂β₁ (antibody 1936) and then reprobed with antibodies against the α_IIb subunit of integrin α_IIbβ₃ and against the GPIbα subunit of the GPIb-V-IX complex, as indicated on the right. The positions of molecular weight markers are reported on the left.