Two amperometric biosensors based on polyvinylferrocenium (PVF+) and platinum deposited PVF+ were constructed for lysine detection. The method for modification of electrodes relied on the electrooxidation of polyvinylferrocene in methylene chloride containing tetrabutylammonium perchlorate (TBAP) at +0.70 V and electrodeposition of platinum particles on the surface of PVF+ClO4- at -0.15 V vs. Ag/AgCl. Lysine oxidase (LyOx) enzyme was immobilized by ion exchange process into modified electrodes. The response of the resulted PVF(+)LyOx(-) and PVF(+)LyOx(-)/Pt biosensors determined by oxidation of enzymatically produced H2O2 at 0.60 V vs. Ag/AgCl. The influence of experimental conditions such as applied potential, pH, temperature on the biosensor responses were examined and analytical performance characteristics of the biosensors were determined. The experimental results of two biosensors compared with each other. The results showed that PVF(+)LyOx(-)/Pt biosensor had a linear range up to 3.0 mM whereas for PVF(+)LyOx(-) biosensor that range was linear up to 1.3 mM. The detection limits of PVF(+)LyOx(-)/Pt and PVF(+)LyOx(-) biosensors were determined to be about 6.5 x 10(-4) and 5.0 x 10(-4) mM, respectively. The response time of sensors was less than 30 s. The anti-interference ability and long-term stability of the biosensors were studied. The proposed biosensors were also used to determine lysine in pharmaceutical samples. (C) 2012 Elsevier B.V. All rights reserved.