Determination of apo-metallothionein using adsorptive transfer stripping technique in connection with differential pulse voltammetry

Links between metallothionein (MT), its structure and many biologically important pathways demonstrate the necessity of taking into account of studying the behavior of MT in different well defined mediums, which could help to model conditions in an organism easily. The main aim of this work was to prepare and determine apoMT by adsorptive transfer stripping technique (AdTS) in connection with differential pulse voltammetry (DPV). Particularly, we investigated the electrochemical behavior of MT measured on the surface of hanging mercury drop electrode in the presence of sodium chloride as supporting electrolyte using AdTS DPV with respect to study the effects of MT signals by different concentrations and pH's of the electrolyte. Then, we aimed at utilizing this technique to observe changes in MT which are dependent on using strong chelating compounds ethylendiamine-N, N, N', N'-tetraacetic acid (EDTA). Thanks to ability of EDTA to bind heavy metals from active center of enzymes and regulation proteins, we were able to prepare apoMT; that means, MT without any metal ion bound in its structure. Detection limit of apoMT at very short time of accumulation (t(A) = 120 s) was 3 nM (20 ng/mL and/or 15 fmol in 5 mu L drop; RSD 2-5%) estimated by dilution of the analyzed solution until the signal disappeared. In addition, it was possible to decrease detection limit by extending of time of accumulation of apoMT on the surface of HMDE. We were able to detect 30 pM of apoMT (200 pg/mL and/or 150 amol in 5 mu L drop) at t(A) = 500 s.