A dataset of knee kinematics in healthy, uninjured adults is needed to serve as a reference for comparison when evaluating the effects of injury, surgery, rehabilitation, and age. Most currently available datasets that characterize healthy knee kinematics were developed using conventional motion analysis, known to suffer from skin motion artifact. More accurate kinematics, obtained from bone pins or biplane radiography, have been reported for datasets ranging in size from 5 to 15 knees. The aim of this study was to characterize tibiofemoral kinematics and its variability in a larger sample of healthy adults. Thirty-nine knees were imaged using biplane radiography at 100 images/s during multiple trials of treadmill walking. Multiple gait trials were captured to measure stance and swing-phase knee kinematics. Six degrees-of-freedom kinematics were determined using a validated volumetric model-based tracking process. A bootstrapping technique was used to define average and 90% prediction bands for the kinematics. The average ROM during gait was 7.0 mm, 3.2 mm, and 2.9 mm in anterior/posterior (AP), medial/lateral (ML), and proximal/distal (PD) directions, and 67.3 deg, 11.5 deg, and 3.7 deg in flexion/extension (FE), internal/external (IE), and abduction/adduction (AbAd). Continuous kinematics demonstrated large interknee variability, with 90% prediction bands spanning approximately ±4 mm, ±10 mm, and ±5 mm for ML, AP, and PD translations and ±15 deg, ±10 deg, and ±6 deg in FE, IE, and AbAd. This dataset suggests substantial variability exists in healthy knee kinematics. This study provides a normative database for evaluating knee kinematics in patients who receive conservative or surgical treatment.