Whilst exercise is beneficial to those suffering from low back pain (LBP), spinal shrinkage and vibration during walking may aggravate the condition. This study investigates the effects of spinal unloading, by means of body-weight unloading (BWU), on spinal length and vibration response. Under-arm partial BWU (40% of bodyweight) was performed on 8 healthy males whilst walking on a treadmill for 1 h, and compared to a control condition in the same participants. Motion analysis was used to track four reflective markers attached to the spine between C7 and the lumbar concavity at 100 Hz, in 7 s samples at regular intervals during the walk. A quintic polynomial was fitted to the coordinates in the sagittal plane, and sub-sectioned into three regions: the upper thoracic (UT), lower thoracic (LT) and upper lumbar (UL). The lengths of the curves were analysed in the time and frequency domains. The length of the spine increased by 4 ± 2% (18 mm) during the unloading condition with all regions showing an increase in length. The UL and LT regions lost length in the control condition, thereby exhibiting a significant interaction between unloading and time on region length (both P < 0.05). In addition, compared to the control condition, the frequencies of the length changes were attenuated between 3 and 6 Hz with unloading. Therefore, under-arm BWU facilitates spinal elongation, attenuates the frequency response of the spine in its resonant frequency regime and thus has potential benefits to the LBP population.