Besides their role in hemostasis, platelets are also highly involved in the pathogenesis and progression of cardiovascular diseases. Since important and initial steps of platelet activation and aggregation are regulated by phosphorylation events, a comprehensive study aimed at the characterization of phosphorylation-driven signaling cascades might lead to the identification of new target proteins for clinical research. However, it becomes increasingly evident that only a comprehensive phosphoproteomic approach may help to characterize functional protein networks and their dynamic alteration during physiological and pathophysiological processes in platelets. In this review, we discuss current methodologies in phosphoproteome research including their potentials as well as limitations, from sample preparation to classical approaches like radiolabeling and state-of-the-art mass spectrometry techniques.