Metallothioneins (MTs) are involved in heavy metal detoxification in a wide range of living organisms. Currently, it is well known that MTs play substantial role in many pathophysiological processes, including carcinogenesis, and they can serve as diagnostic biomarkers. In order to increase the applicability of MT in cancer diagnostics, an easy-to-use and rapid method for its detection is required. Hence, the aim of this study was to develop a fully automated and high-throughput assay for the estimation of MT levels. Here, we report the optimal conditions for the isolation of MTs from rabbit liver and their characterization using MALDI-TOF MS. In addition, we described a two-step assay, which started with an isolation of the protein using functionalized paramagnetic particles and finished with their electrochemical analysis. The designed easy-to-use, cost-effective, error-free and fully automated procedure for the isolation of MT coupled with a simple analytical detection method can provide a prototype for the construction of a diagnostic instrument, which would be appropriate for the monitoring of carcinogenesis or MT-related chemoresistance of tumors.

Department of Biochemistry Faculty of Science Charles University Albertov 2030 CZ 128 40 Prague 2 Czech Republic

Department of Chemistry and Biochemistry Mendel University in Brno Zemedelska 1 CZ 613 00 Brno Czech Republic; Central European Institute of Technology Brno University of Technology Purkynova 123 CZ 612 00 Brno Czech Republic

Department of Paediatric Haematology and Oncology 2nd Faculty of Medicine Charles University and University Hospital Motol 5 Uvalu 84 CZ 150 06 Prague 5 Czech Republic

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