Purpose Development of adjacent segment degeneration following anterior cervical decompression and fusion (ACDF) is still controversial, as adjacent-level kinematics is poorly understood. This study reports preliminary data from a high-accuracy 3D analysis technique developed for in vivo cervical kinematics. Methods From nine cervical spondylosis patients, four underwent single-level ACDF, and five underwent two-level ACDF using cylindrical titanium cage implant(s). Pre-and post-surgical CT scans were taken in flexion, neutral and extended positions, allowing us to compute segmental ranges of motion for rotation and translation, and 3D disc-height distributions. Differences in segmental motions and disc-height between fused and adjacent levels were analyzed with a Wilcoxon signed-rank test. Results are presented as mean ± SEM. Results The flexion/extension angular-ROM at the fusion level decreased after surgery (7.46 ± 1.17°vs. 3.14 ± 0.56°, p \ 0.003). The flexion/extension angular-ROM at one caudal adjacent level to the fusion level (3.97 ± 1.29°) tended to be greater post-operatively (6.11 ± 1.44°, p = 0.074). Translation in the anterior-posterior direction during flexion/ extension at the fusion level decreased after surgery (1.22 ± 0.20 mm vs. 0.32 ± 0.11 mm, p \ 0.01). No differences were found in adjacent-level disc heights between both study time-points. Conclusions This study showed increased segmental motion in flexion/extension angular-ROM at one level adjacent to ACDF. However, increases in the rotational angular-ROM were not statistically significant when cranial/caudal adjacent levels were analyzed separately. This preliminary study highlighted the capabilities of a 3D-kinematic analysis method to detect subtle changes in kinematics and disc height at the adjacent levels to ACDF. Thus, reliable evidence related to ACDF's influence on adjacent-level cervical kinematics can be collected.