Flow cytometry analysis of intracellular VASP phosphorylation for the assessment of activating and inhibitory signal transduction pathways in human platelets - Definition and detection of ticlopidine/clopidogrel effects

Increased platelet adhesion or aggregation are key events in the pathogenes is of cardiovascular diseases. Exact determination of the platelet activati on state is essential to recognize, prevent, and treat cardiovascular compl ications due ra platelet dysfunction. initial phases of platelet activation and inhibition are characterized by phosphorylation of specific intracellu lar proteins. However, methodological problems often prevent analysis of pl atelet protein phosphorylation under clinical conditions. A novel flow cyto metry-based method using a phosphorylation-specific antibody was developed for fast and easy quantification of the phosphorylation state of a specific intracellular platelet protein. This method was used to analyze various pl atelet receptors and their intracellular signaling which may be impaired in genetic or acquired disorders or altered due to therapeutic interventions. In a first clinical application, the inhibitory effects of ticlopidine and clopidogrel on the platelet P2Y(AC) ADP receptor were monitored.