The Osprey is operated by a four-person crew consisting of two pilots and two flight engineers. It is utilized for various missions including combat, combat support, combat service support, and Special Operations. The main function of the Osprey is to transport troops, equipment, and supplies from assault ships and land bases in an amphibious manner. The Air Force and U.S. Special Operations Command use the CV-22 for Spec Ops, while the Navy utilizes the HV-22 for Combat Search and Rescue and fleet logistics support. The Marines are also replacing their aging fleet of CH-46E Sea Knight and CH-53D Sea Stallion medium-lift helicopters with the Osprey. The MV-22 will be integrated into all Marine Corps medium-lift active duty and reserve tactical squadrons, the medium-lift training squadron (FRS), and the executive support squadron (HMX).

The Osprey has been deployed globally for a variety of missions such as combat operations, humanitarian efforts, peacekeeping, and regional stability initiatives. It has been utilized in conflict zones including Afghanistan, Iraq, Syria, Libya, parts of Africa, and the Asia-Pacific region for military exercises and other purposes. During missions, the Osprey primarily operates in fixed-wing mode, reducing wear and tear and operational costs. It is also designed to fly at higher altitudes than traditional helicopters, providing longer line-of-sight communications and stronger command and control for the crew.