beta-galactosidase was successfully immobilized onto chitosan (CS)/polyvinyl alcohol (PVA) blend nanofibers through electrospinning. The effects of both polymer solution parameters and the electrospinning process conditions on the morphology of nanofibers and enzyme activity were studied. FESEM results showed that both morphology and diameter of the nanofibers were significantly influenced by the studied factors. The nanofiber mats obtained from CS/PVA at 50: 50 mass ratio and conditions of 20 kV and 0.5 ml/h were selected as the optimized nanofibers, with the smallest diameter and most homogenous structure. On the other hand, catalytic activity of the beta-galactosidase-immobilized nanofibers (Enzyme-NF) was not significantly changed with these factors. The catalytic activity of Enzyme-NF was 57.03% of free enzyme activity at pH 6.8 and 4 degrees C. Furthermore, it showed more temperature stability compared to free beta-galactosidase at 50 degrees C. During 28 days of storage, Enzyme-NF samples retained 77% and 42% of its initial activity in 4 degrees C and 25 degrees C, respectively.