Synthetic organic polymers are commonly used to facilitate challenging solid-liquid separations such as biosludge dewatering. However, there is interest in reducing the use of polymers due to their toxicity and synthetic sourcing. Surfactants and proteins have shown potential to enhance sludge dewaterability but little is known about the properties and/or mechanism(s) that promote this enhancement. In this study, synthetic polymers, surfactants and proteins were investigated to evaluate whether surface properties such as charge, surfactant activity and hydrophobicity, play a role in how these conditioners affect biosludge dewatering. Capillary suction time (CST), dry solids content, filtrate rate and filtrate solids content were used to assess dewaterability. Results show that surface charge determines the potential of conditioners. The effect of charge was greater for surfactants and proteins than for polymers. In contrast with previous reports, surfactant activity negatively affected the dewaterability of biosludge. Cationic conditioners, regardless of the group improved biosludge dewaterability. However, the dose of cationic proteins is still high compared to currently used synthetic polymers (e.g. protamine is 0.1 g/g TSS vs. synthetic polymer 0.03 g/g TSS). Our results suggest that there is potential for using proteins to improve biosludge dewaterability but a further reduction in protein dose and/or an increase in the protein's efficiency as a conditioner is needed.