Lipid raft disruption by MβCD impairs CLEC-2 signaling. (A) Platelets were pretreated with indicated concentrations of MβCD and stimulated with 30nM rhodocytin or 0.1 U/mL thrombin. (Left) Representative aggregation traces are shown for control and MβCD-treated platelets. Addition of agonist is indicated by an arrowhead. The aggregation traces have been staggered. (Right) The percentage of transmittance of platelets pretreated with decreasing concentrations of MβCD-treated platelets stimulated with rhodocytin or thrombin was measured after 5 minutes. Data represent the mean and standard error of 3 independent experiments. (B) CLEC-2 was immunoprecipitated from platelets pretreated with indicated concentrations of MβCD and stimulated with vehicle or 30nM rhodocytin and blotted with an antiphosphotyrosine antibody. Membranes were subsequently stripped and reblotted with anti–CLEC-2. Data are representative of 3 independent experiments.