Improving walking ability is an important aim in rehabilitation after stroke. A relatively new method to improve walking ability is robot assisted treadmill training, which assists the lower limbs by providing a quasi normal walking pattern. Whether this training method has an effect on the muscle activity is unknown in patients with hemiplegia due to stroke. PURPOSE: To evaluate muscle activity of individuals with stroke during robotic treadmill walking compared to overground walking. METHODS: Ten subjects (6 men, age 55±11 yrs) with chronic stroke (mean 65 wks) with either a left (n=2) or a right (n=8) hemiparesis and a FAC score of 5 (able to walk without assistance) participated. During robotic walking (RW), subjects walked in a robotic walking device (Lokomat) with minimal support (minimal body weight support and guidance force) and a walking speed of 2.2 km/h. During overground walking (OW), subjects walked without assistance at their preferred walking speed. EMG data of ten gait cycles were collected from 7 leg muscles, high pass filtered (20 Hz), rectified, low pass filtered (4 Hz), and normalized (%maximal EMG amplitude). Differences in amplitude were investigated by a multifactor ANOVA and an additional t-test. RESULTS: Average EMG amplitude in the RW condition was significantly (p<0.05) lower than in the OW condition for the rectus femoris, gluteus maximus and tibialis anterior muscles. Significant interaction effects indicated a higher activity of the paretic semitendinosus and gastrocnemius muscle during RW compared to OW. The lower mean activity during RW than during OW can be explained by the stabilization of the gait orthosis reducing the co activation of the lower-limb muscles. Furthermore, the support of the gait orthosis reduces the effort of the legs during walking which reduces the muscle activity. CONCLUSION: Overall, muscle activity was lower during RW than during OW. The interaction effects, however, suggest that although the overall activity of the muscles was lower during RW, the activity of the paretic semitendinosus, gastrocnemius and tibialis anterior muscles had a lower decrease or even an increase during RW, supporting the use of robot assisted treadmill training in rehabilitation after stroke.