Peptidoglycan (PGN), a component of bacterial cell walls and belonging to "Microbe-Associated Molecular Patterns" (MAMP) triggers host reactions contributing to the pathophysiology of infectious disease. Host cell responses to PGN exposure include apoptosis. Bacterial infections may result in activation of blood platelets and thrombocytopenia. The present study explored, whether HPLC-purified fractions of PGNs from Staphylococcus aureus 113 triggers apoptosis of platelets. To this end platelets were exposed to PGN fractions and annexin-V binding determined to depict cell membrane scrambling, DiOC6 fluorescence to estimate depolarization of mitochondrial potential, Fluo-3AM staining for intracellular Ca²⁺ activity ([Ca²⁺]_i) and immunofluorescence to quantify protein abundance of active caspase-3. As a result, a 30 min exposure to PGN monomeric fraction (>=50 ng/ml) was followed by annexin-V binding, increase of [Ca²⁺]_i, mitochondrial depolarization, caspase-3 activation and CD41/61 upregulation. The annexin-V binding was significantly blunted by anti-TLR-2 antibodies, in the absence of extracellular Ca²⁺ and in the presence of pancaspase inhibitor. In conclusion, PGN triggers apoptosis of platelets in activation-dependent manner, characterized by mitochondrial depolarization, caspase-3 activation and cell membrane scrambling.