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Stair climbing under backpack load condition is a challenging task. Understanding muscle activation patterns of lower limb during stair climbing with load furthers our understanding of the factors involved in joint pathology and the effects of treatment. At the same time, stair climbing under backpack load requires adjustments of muscle activations and increases joint moment compared to level walking, which with muscle activation patterns are altered as a result of using an assistive technology, such as a wearable exoskeleton leg for human walking power augmentation. Therefore, the aim of this study was to analyze lower limb muscles during stair climbing under different backpack load. Nine healthy volunteers ascended a four-step staircase at different backpack load (0 kg, 10 kg, 20 kg, 30 kg). Electromyographic (EMG) signals were recorded from four lower limb muscles (gastrocnemius, tibialis anterior, hamstring, rectus femoris). The results showed that muscle activation amplitudes of lower limb increase with increasing load during stair climbing, the maximum RMS of gastrocnemius are greater than tibialis anterior, hamstring and rectus femoris whether stair climbing or level walking under the same load condition. However, the maximum RMS of hamstring are smaller than gastrocnemius, tibialis anterior and rectus femoris. The study of muscle activation under different backpack load during stair climbing can be used to design biomechanism and explore intelligent control based on EMG for a wearable exoskeleton leg for human walking power augmentation.