The effect of ratio between autoflocculating and target microalgae in bio-flocculation was studied with emphasis on the recovery, sedimentation rate and energy demand for harvesting the target microalgae. When the autoflocculating microalgae Ettlia texensis, Ankistrodesmus falcatus and Scenedesmus obliquus were added to Chlorella vulgaris at a ratio of 0.25, the recovery of C. vulgaris increased from 25% to, respectively, 40%, 36% and 31%. The sedimentation rate increased as well. Addition of Tetraselmis suecica to Neochloris oleoabundans at a ratio of 0.25 increased the recovery from 40% to 50%. Application of bio-flocculation at a ratio of 0.25, followed by centrifugation reduces the energy demand for harvesting of the target microalgae from 13.8 MJ kgDW(-1) if only centrifugation is used to 1.83, 1.81, 1.53 and 1.34 MJ kgDW(-1), respectively, using T. suecica, E. texensis, A. falcatus and S. obliquus and 3h sedimentation before centrifugation.