Blood flow restriction (BFR) during low-intensity exercise has been known to be a potent procedure to alter metabolic and oxygen environments in working muscles. Moreover, the use of BFR during inter-set rest periods of repeated sprint exercise has been recently suggested to be a potent procedure for improving training adaptations. The present study was designed to determine the effect of repeated sprint exercise with post-exercise BFR (BFR during rest periods between sprints) on muscle oxygenation in working muscles. Eleven healthy males performed two different conditions on different days: either repeated sprint exercise with BFR during rest periods between sets (BFR condition) or without BFR (CON condition). A repeated sprint exercise consisted of three sets of 3 × 6-s maximal sprints (pedaling) with 24s rest periods between sprints and 5 min rest periods between sets. In BFR condition, two min of BFR (100-120 mmHg) for both legs was conducted between sets. During the exercise, power output and arterial oxygen saturation (SpO2 ) were evaluated. Muscle oxygenation for the vastus lateralis muscle, exercise-induced changes in muscle blood flow, and muscle oxygen consumption were measured. During BFR between sets, BFR condition presented significantly higher deoxygenated hemoglobin + myoglobin (p < 0.01) and lower tissue saturation index (p < 0.01) than those in CON condition. However, exercise-induced blood lactate elevation and reduction of blood pH did not differ significantly between the conditions. Furthermore, power output throughout nine sprints did not differ significantly between the two conditions. In conclusion, repeated sprint exercise with post-exercise BFR augmented muscle deoxygenation and local hypoxia, without interfering power output.