The development of effective treatment and reconstruction procedures for injuries to the soft tissues around the acromioclavicular (AC) joint relies on a comprehensive understanding of overall joint function. The objective of this study was to determine the magnitude and direction of the in situ forces in the AC capsular and coracoclavicular ligaments as well as the resulting joint kinematics during application of three external loading conditions while allowing three degree-of-freedom joint motion. A robotic/universal force-moment sensor testing system was utilized to determine the in situ forces in the soft tissue structures and the resulting joint kinematics. The clavicle translated 5.1+/-2.0, 5.6+/-2.2, and 4.2+/-1.9 mm during application of a 70 N load in the anterior, posterior, and superior directions, respectively, representing almost a 50% increase over previous studies using similar load magnitudes. In response to an anterior load, the magnitude of in situ force in the superior AC ligament (35+/-18 N) was found to be greater (p<0.05) than the force in the trapezoid and conoid ligaments. In contrast, the magnitude of in situ force in the conoid (49+/-22 N) was significantly greater (p<0.05) than all other ligaments in response to a superior load. Additionally, the directions of the force vector representing the conoid and trapezoid were different, being located in opposing quadrants of the posterior axis of the scapula with this loading condition. Our data suggest that the kinematic constraints placed on the AC joint during loading affect the resulting joint motion and that the magnitude and direction of force in each ligament are affected by the coupled motions that occur. Based on the differences in magnitude and direction of the in situ force in the coracoclavicular ligaments with each loading condition, surgical procedures should reconstruct these ligaments in a more anatomical manner or treat them separately to prevent joint degeneration.