While inclined walking is a frequent daily activity, muscle forces during this activity have rarely been examined. Musculoskeletal models are commonly used to estimate internal forces in healthy populations, but these require a priori validation. The aim of this study was to compare estimated muscle activity using a musculoskeletal model with measured EMG data during inclined walking. Ten healthy male participants walked at different inclinations of 0°, ± 6°, ± 12°, and ± 18° on a ramp equipped with 2 force plates. Kinematics, kinetics, and muscle activity of the musculus (m.) biceps femoris, m. rectus femoris, m. vastus lateralis, m. tibialis anterior, and m. gastrocnemius lateralis were recorded. Agreement between estimated and measured muscle activity was determined via correlation coefficients, mean absolute errors, and trend analysis. Correlation coefficients between estimated and measured muscle activity for approximately 69% of the conditions were above 0.7. Mean absolute errors were rather high with only approximately 38% being ≤ 30%. Trend analysis revealed similar estimated and measured muscle activities for all muscles and tasks (uphill and downhill walking), except m. tibialis anterior during uphill walking. This model can be used for further analysis in similar groups of participants.