The influence of in-feed lactoferrin (Lf) on bird production, intestinal microbiota, mucosal immune system and gut microarchitecture was assessed in male Cobb 500 broiler chickens. Birds were given one of four diets from day of hatch: Control (basal diet with no additives), ZnB (basal diet + 50 mg/kg zinc bacitracin), Lf 250 mg/kg (basal diet + 250 mg/kg Lf) and Lf 500 mg/kg (basal diet + 500 mg/kg Lf); n = 24 birds/treatment. An apparent metabolisable energy study was performed between d 25-32. Lf did not affect growth rate or feed conversion in the period 0-21 d of age, nor performance or energy metabolism during the 7 d metabolism experiment which commenced at 25 d of age.The profiles of caecal microbial communities were significantly different in birds given ZnB compared with birds given a diet with no additives, or supplemented with 250 mg/kg Lf. Birds given 250 mg/kg Lf also had a different microbial profile compared with birds given 500 mg/kg Lf. In comparison to control birds, Lf treated birds showed some differences in the T cell proportions in caecal tonsil and spleen. No differences in ileal villus height, crypt depth or goblet cell proportions were observed amongst dietary treatments. Whilst Lf had little effect on the measured parameters, the use of an integrated approach to study the influence of novel feed additives may facilitate a greater understanding of the relationships between nutrition, gut health and bird performance.