The posterior cruciate ligament (PCL) functions as the primary restraint to posterior tibial movement and the stabilizer for axial rotation of the knee joint. It is, therefore, important to know the mechanical characteristics of the PCL. The objective of this study was to evaluate the in vivo characteristics of the three-dimensional (3D) deformation and orientation of the PCL during knee flexion using magnetic resonance imaging (MRI). Ten normal knees were visualized using MRI at full extension (FE), 10°, 20°, 30° and 60° of static knee flexion. The length, strain, elevation and deviation of the PCL were measured using the 3D models of the femur, tibia and PCL reconstructed from the MR images. The PCL length was measured in three portions (Medial: M, Central: C, Lateral: L) to consider its 3D shape. Tendencies of the PCL elongation and strain during knee flexion were different among these 3 portions. The PCL length in M portion was shortest in 10° or 20° and extended in FE and 60°. The PCL length in L portion, on the other hand, extended from FE to 60°. These 3D deformation characteristics were consisted with the changes in the elevation and deviation of the PCL during knee flexion.