Non-porous poly(2-hydroxyethyl methacrylate) [p(HEMA)] nanoparticles were prepared by surfactant free emulsion polymerization. The p(HEMA) nanoparticles was about 200 nm diameter, spherical form, and non-porous. Reactive Red 120 (RR 120) was covalently attached to the p(HEMA) nanoparticles and Ni(II) ions were incorporated to attach dye molecules. Urease was immobilized onto RR120-Ni(II) attached p(HEMA) nanoparticles via adsorption. The maximum urease adsorption capacity of RR120-Ni(II) attached p(HEMA) nanoparticles was 480.01 mg g−1 nanoparticles at pH 7.0 in phosphate buffer. It was observed that urease could be repeatedly adsorbed and desorbed without significant loss in adsorption amount. Km values were 21.50 and 34.06 mM for the free and adsorbed enzyme. The Vmax values were 4 U for the free enzyme and 3.3 U for the adsorbed enzyme. The optimum pH was 25 mM pH 7 phosphate buffer for free and adsorbed enzyme. The optimum temperature was determined at 35°C and 55°C for the free and adsorbed enzyme, respectively. These findings show considerable promise for this material as an adsorption matrix in biotechnological applications. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014, 131, 39757.