Increased levels of light, moderate and vigorous physical activity (PA) are positively associated with health benefits. Therefore, sensor-based human activity recognition can identify different types and levels of PA. In this paper, we propose a two-layer locomotion recognition method using dynamic time warping applied to inertial sensor data. Based on a video-validated dataset (ADAPT), which included inertial sensor data recorded at the lower back (L5 position) during an unsupervised task-based free-living protocol, the recognition algorithm was developed, validated and tested. As a first step, we focused on the identification of locomotion activities walking, ascending and descending stairs. These activities are difficult to differentiate due to a high similarity. The results showed that walking could be recognized with a sensitivity of 88% and a specificity of 89%. Specificity for stair climbing was higher compared to walking, but sensitivity was noticeably decreased. In most cases of misclassification, stair climbing was falsely detected as walking, with only 0.2–5% not assigned to any of the chosen types of locomotion. Our results demonstrate a promising approach to recognize and differentiate human locomotion within a variety of daily activities.