Poly vinyl acetate grafted calcium alginate hydrogel microspheres were prepared with bovine serum albumin (BSA) as molecular template. The microspheres exhibited homogeneous and core-shell structure according to different preparation strategy. The rebinding and swelling property of microspheres showed responsiveness toward ionic strength, temperature, and pH. It was found the highest separation factor of 1.85 and imprinting efficiency of 1.75 when the Ca2+ ionic strength was 1.25 and 0.34 mol/kg, respectively. The separation factor was found decreased as temperature grew from 29 to 45 degrees C while the imprinting efficiency reached a peak value at about 37 degrees C. Separation factor of BSA imprinted microspheres at different pH was also recorded and two peaks were found, which were considered to be caused by the similar swollen state due to ionic and covalent crosslinking structure of modified microspheres. The recognition responsiveness is suggested to be influenced by environmental effects due to the changing in imprints' configuration at different swollen states. (c) 2012 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013