Background: Most patients with isolated posterior cruciate ligament (PCL) injuries have minimal symptoms, and nonoperative treatment is recommended. However, over time, these patients can develop significant degenerative changes in their knees. Historically, PCL laxity is graded by nonweightbearing anteroposterior measuring techniques that do not reproduce the true, dynamic weightbearing conditions in the injured knee. The purpose of this study was to determine the patholaxity in patients with isolated PCL deficiency during functional weightbearing activities (running, walking, and stair ascent). Hypothesis: Patients with unilateral, isolated PCL deficiency will demonstrate dynamic anteroposterior and rotational instability in their affected knees during functional activities of level running and stair ascent compared with their unaffected, contralateral knees. Study Design: Controlled laboratory study. Methods: Nine asymptomatic patients with isolated grade II PCL injury underwent Dynamic Stereo X-Ray (DSX) of both knees during level running and stair ascent. Three-dimensional reconstructions of the patients’ bilateral distal femurs and proximal tibias were created from high-resolution computed tomography (CT) scans. Three-dimensional joint kinematics were determined using a model-based tracking approach to align the radiographic images with CT-derived bone models. The resulting tibiofemoral rotations and translations for the PCL-deficient and PCL-intact knees were then compared. Results: During level running, the tibia of the PCL-deficient knee was approximately 2 mm posteriorly subluxated and had an anterior velocity relative to the femur approximately 40 mm/s greater than the contralateral, uninjured knee; however, this was only during the swing phase. No significant differences were found during the stance phase of running. During stair ascent, the tibia of the PCL-deficient knee was approximately 4 mm posteriorly subluxated compared with the intact limb during the terminal swing phase and early stance phase. Between foot strike and the time of peak ground-reaction force (GRF), the tibia of the PCL-deficient knee translated anteriorly relative to the femur with velocities 3 to 4 times greater than in the intact limb. Level walking was also evaluated in 3 patients, but no differences were seen, and it was not tested in the remaining 6 patients. Conclusion: Changes in knee kinematics due to isolated PCL injuries were highly activity dependent. During running, small differences were identified only during the swing phase when the knee was unloaded. However, during stair ascent, significant differences extended from the late swing into early stance phase. During the swing phase of stair ascent, the tibia in the PCL-deficient joint subluxated posteriorly. Then, as load was transferred to the ascending limb, the tibia reduced anteriorly with high velocity relative to the femur. The resulting shear motion may expose the loaded joint to abnormal and potentially damaging forces. Clinical Relevance: During functional activities, patients with isolated PCL injuries experience significant knee instability that cannot be identified by standard nonweightbearing static laxity measurements. The finding that different activities create different degrees of instability may have important implications for rehabilitation and activity limitations for PCL-deficient individuals.