Three production trials and one nitrogen balance trial were conducted with Lohmann Brown hens to determine the requirement for and effects of an excess of isoleucine in layers at different ages (24 to 32 and 46 to 54 wk of age). The trials were designed as dose-response studies where isoleucine-deficient basal rations with 11.4 MJ metabolizable energy per kilogram were supplemented with varying amounts of L-isoleucine. In the production trials, dietary isoleucine concentrations ranged from 0.37 to 1.05%. In the three production trials, maximum daily egg mass was achieved at dietary isoleucine concentrations of between 0.39 and 0.75% (25 to 32 wk of age, daily egg mass 53 g), 0.40 and 0.57% (24 to 32 wk of age, daily egg mass 57 g), and 0.40 and 0.81% (46 to 54 wk of age, daily egg mass 56 g). The corresponding ranges of daily isoleucine intakes were 412 to 770 mg, 436 to 624 mg, and 431 to 874 mg. In the nitrogen balance trial, maximum total nitrogen retention was achieved at dietary isoleucine concentrations of between 0.43 and 0.57%. Dietary isoleucine concentrations higher than 0.8% caused a reduction in hen BW. Dietary isoleucine concentrations higher than 1.0% additionally caused a reduction in the daily egg mass. The study thus shows that the margin between requirement and excess of isoleucine is narrow in laying hens.