In vitro and in vivo targeting of scFvSCE5 in mouse platelets. (A) Flow cytometric detection of scFv-CD39 constructs using an Alexa Fluor-488–coupled anti–Penta-His antibody that binds to the constructs’ 6xHis-tags. Representative histograms showing binding of targeted or nontargeted CD39 constructs as assessed by a secondary anti–Penta-His Alexa Fluor-488 antibody. Nonactivated and activated platelets were differentiated by gating with a PE-labeled anti–P-selectin antibody. (B) Intravital microscopy of delayed binding of scFvSCE5 to adhering platelets. Representative images of the formation of a platelet layer induced by ferric chloride injury in the mesenteric artery. ScFvSCE5 binding was detected by an anti–Penta-His Alexa Fluor-488 antibody. Time point 0 depicts time after 4 minutes of ferric chloride–induced injury and 1 minute after flushing with saline when platelets began to attach to the injured site.
