Talin1(L325R) binds to integrins but is defective in agonist-induced platelet integrin activation. (A) Affinity chromatography with the use of recombinant β1, β3, and αIIb cytoplasmic domains and platelet lysates from Tln1^(wt/fl),Cre⁺ (indicated as wt) or Tln1^(L325R/fl),Cre⁺ (indicated as L325R) mice. Bound talin protein was visualized by Western blotting with anti-talin 8d4 antibody, and equal loading of integrins was verified by Coomassie stain. Densitometric quantitation of talin bound was normalized to integrin loading. The value shown indicates the amount of talin1(L325R) bound to the integrin relative to talin1 (wt). Data are representative of 2 independent experiments. (B) Specific binding of fluorescein isothiocyanate-fibrinogen to platelets was measured by the use of flow cytometry by subtracting the nonspecific fibrinogen binding that occurred in the presence of 5mM ethylenediaminetetraacetic acid in each condition. *P < .05, **P < .005. (C) Activation of β1 integrin was assessed by measuring the binding of the conformation-sensitive antibody 9EG7 relative to binding of the conformation-insensitive β1 integrin antibody HMβ1-1. **P < .005.