The levels of platelet-derived extracellular vesicles (pEVs) have been reported as elevated in acute ischemic stroke (IS). However, the results of studies remain equivocal. This prospective, case-control study included 168 patients with IS, 63 matched disease controls (DC), and 21 healthy controls (HC). Total pEVs concentration, the concentration of phosphatidylserine-positive pEVs (PS⁺pEVs), the percentage of PS⁺pEVs (%PS⁺pEVs) and the concentration of pEVs with expression of CD62P⁺, CD40L⁺, CD31⁺, and active form of GPIIb/IIIa receptor (PAC-1⁺) were assessed on days 1, 3, 10, and 90 with the Apogee A50-Micro flow cytometer. The concentrations of pEVs, PS⁺pEVs, and %PS⁺pEVs were significantly higher after IS vs. HC (p < 0.001). PS⁺pEVs were higher after stroke vs. controls (p < 0.01). The concentrations of pEVs with expression of studied molecules were higher on D1 and D3 after stroke vs. controls. The concentration of pEVs after platelet stimulation with ADP was significantly diminished on D3. IS most notably affects the phenotype of pEVs with a limited effect on the number of pEVs. Ischemic stroke moderately disturbs platelet microvesiculation, most notably in the acute phase, affecting the phenotype of pEVs, with a limited impact on the number of pEVs.