Muscle disuse results in the loss of muscular strength and size, due to an imbalance between protein synthesis (MPS) and breakdown (MPB). Protein ingestion stimulates MPS, although it is not established if protein is able to attenuate muscle loss with immobilization (IM) or influence the recovery consisting of ambulatory movement followed by resistance training (RT). Thirty men (49.9±0.6 years) underwent 14 days of unilateral leg IM, 14 days of ambulatory recovery (AR) and a further 6 RT sessions over 14 days. Participants were randomized to consume an additional 20g of dairy protein or placebo with a meal during the intervention. Isometric knee extension strength was reduced following IM (-24.7 ± 2.7 %), partially recovered with AR (-8.6 ± 2.6%) and fully recovered after RT (-0.6 ± 3.4 %), with no effect of supplementation. Thigh muscle cross sectional area decreased with IM (-4.1 ± 0.5 %), partially recovered with AR (-2.1 ± 0.5 %), and increased above baseline with RT (+2.2 ± 0.5 %), with no treatment effect. Myofibrillar MPS, measured using deuterated water, was unaltered by IM, with no effect of protein. During AR MPS was increased only with protein supplementation. Protein supplementation did not attenuate the loss of muscle size and function with disuse or potentiate recovery, but enhanced myofibrillar MPS during AR.