Human Skeletal Muscle Myoblasts usually begin to form multinucleated myotubes when the culture reaches 50-70% confluence.

Myotubes are formed by the fusion of several to many myoblasts. Addition of differentiation medium, when the culture reaches approximately 50% confluence, will promote the maturation of myotubes. The procedure linked below describes how to differentiate the human skeletal muscle myoblasts (HSMM) using a differentiation medium, also referred to as a fusion medium.