Paramagnetic antibody-modified microparticles coupled with voltammetry as a tool for isolation and detection of metallothionen as a bioindicator of metal pollution

Abstract

Low-molecular mass proteins rich in cysteines called metallothioneins (MT) can be considered as markers for the pollution of the environment by metals. Here, we report on suggestion for an automated procedure for the isolation of MT followed by voltammetric analysis. Primarily, we optimized the automated detection of MT using an electrochemical analyser. It was found that the most sensitive and repeatable analyses are obtained at a temperature of 4 °C for the supporting electrolyte. Further, we optimized experimental conditions for the isolation of MT by using antibody-linked paramagnetic microparticles. Under the optimal conditions (4 h long interaction between the microparticles and MT), the microparticles were tested on isolation of various amounts of MT. The lowest isolated amount of MT by antibody-linked paramagnetic microparticles was 5 μg ml⁻¹ of MT (50 ng). The automated procedure of MT isolation was further tested on isolation of MT from guppy fish (Poecilia reticulata) treated with silver(I) ions (50 μM AgNO₃). The whole process lasted less than five hours and was fully automated. We attempted to correlate these results with the standard method for MT isolation. The correlation coefficient is 0.9901, which confirms that results are in good agreement. Moreover, the concentration of silver ions in tissues of fish treated with Ag(I) ions was determined by high performance liquid chromatography with electrochemical detection.