Skorobogat'ko Ov

A new approach to the construction of potentiometric immunosensors

An electrochemical immunosensor based on a new detection principle was developed. Laccase, which is able to catalyse the electroreduction of oxygen via the direct (mediatorless) mechanism was used as an enzyme label. The new detection method does not require the presence of an electrochemically active mediator, and the reaction substrates are atmospheric oxygen and electrons, the latter being taken by the active site of the enzyme label directly from the electrode. The formation of the complex between laccase-labelled antibody and antigen on the electrode surface resulted in a considerable (more than 300 mV) shift of the electrode potential. The rate of the increase of the electrode potential was inversely proportional to the concentration of the free antigen in the sample. The non-specific adsorption of conjugate and other proteins on the electrode could be eliminated by using a polyethylenimine-based polymer on the electrode surface. Insulin was used as a model analyte. The sensitivity limit for this antigen was approximately 3 micrograms ml-1.

Laccase as a New Enzymatic Label for Enzyme Immunoassay

Abstract A new immunochemical reagent is suggested containing as an enzyme marker laccase obtained from the cultural liquid of basidial fungi Coriolus hirsutus. The feasibility of immuno-laccase conjugates in different versions of immunoanalysis (sandwich, competitive and indirect enzyme immunoassay) was demonstrated. The assay based on antibodvlaccase conjugates is simpler than that employing antibody-peroxidase conjugates, since in the former case air oxygen is used as the second substrate of the enzymatic reaction.