In this study, a fungal and two yeast -galactosidases were immobilized using alginate and chitosan. The biochemical parameters and lactose hydrolysis abilities of immobilized enzymes were analyzed. The pH optima of immobilized fungal -galactosidases shifted to more acidic pH compared to free enzyme. Remarkably, the optimal temperature of chitosan-entrapped yeast enzyme, Maxilact, increased to 60 degrees C, which is significantly higher than that of the free Maxilact (40 degrees C) and other immobilized forms. Chitosan-immobilized A. oryzae -galactosidase showed improved lactose hydrolysis (95.7%) from milk, compared to the free enzyme (82.7%) in 12h. Chitosan-immobilized Maxilact was the most efficient in lactose removal from milk (100% lactose hydrolysis in 2h). The immobilized lactases displayed excellent reusability, and chitosan-immobilized Maxilact hydrolyzed >95% lactose in milk after five reuses. Compared to free enzymes, the immobilized enzymes are more suitable for cost-effective industrial production of low-lactose milk due to improved thermal activity, lactose hydrolysis efficiencies, and reusability.