The objective of anaerobic digester start-up is to achieve steady-state operation and the required reduction performance in the shortest possible time. The present study aims to assess a novel strategy to start-up pilot scale mesophilic anaerobic digester with internal inoculums; the operational parameters such as feed concentration and operating temperature were also evaluated. Three pilot scale anaerobic digester systems were designed and manufactured. They were equipped with all necessary instruments for measurements, operation and control. Each system consists of a feed tank, a reactor (anaerobic digester), a gas storage tank, and a displaced water collecting tank. The digester start-up was achieved successfully and smoothly without any operational problems. The feed concentration had minor effects on the performance of the digester start-up while the operating temperature has approximately no effect.