Myokines released by skeletal muscle in response to contraction may contribute to the health-promoting effects of exercise. Previous studies with cultured rodent and human myotubes have revealed highly complex patterns of myokine secretion. However, the commonalities and differences in the secretory response of the different cell models have not been explored, limiting the interpretation of these results. In the present study, we performed a comprehensive analysis of contraction-regulated secretomes using the most commonly used skeletal muscle cell models.