Several methods for protein architecture are reported i.e.: layer-by-layer technique, electrodeposition and Langmuir-Blodgett (LB) technique.

LB technology is known as a useful method for the formation of well-oriented film of bio/organic material on a solid substrate, because it can control the degree of order and packing density of protein and especially minimize the loss of protein activity [2]. The structures and moieties of this type of layers with embedded immobilized enzymes are of great interest because of their biosensor applications.

Another common method for immobilization enzymes is electropolymerization and electrodeposition. Electropolymerization methods have been extensively used to create immobilized enzyme electrodes with conducting polymers. The polymer forms a self-assembled monolayer by electrostatically adsorbing to a charged surface.

Continuing systematic investigation in fabrication of protein – conducting material sensitive layers [3], we reported here the results of research directed into the properties of laccase from Cerrena unicolor and tyrosinase from Agaricus bisporus integrated to thin films (LB films, electropolymer layers, Scheme). The immobilized phenoloxidases proved to be an excellent as a bioanalytical tool for monitoring phenolic pollutants. Furthermore, the sensor sensitization can be achieved by interlacing to ordered film conducting amphiphile according successful previous experience (ie carbazole and diphenylamine derivatives) [4]. Conducting polymer - poly[(N-nonylphenoxazine-3,7-diyl-alt-(1,2,3-benzothiadiazole)] admixed into the film is supposed to be a good mediator element in phenol oxidases sensor. The interlaced polymer is expected to facilitate the electron transfer as well enhancing the sensor sensitivity.

                                         Scheme. Immobilization methods of phenoloxidases

[1] B.D. Malhotra, A. Chaubey, S.P. Singh, Anal. Chim. Acta, 2006, 578, 59.

[2] H. G. Choi, B. K. Oh, W. H. Lee, J. W. Choi, Biotechnol. Bioprocess Eng., 2001, 6, 183.

[3] J. Cabaj, J. Soloducho, A. Chyla, J. Bryjak, K. Zynek, Sens. Actuators B, 2009, 136, 425.

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