Iron Ions Released from the Stainless-Steel Anode during High-Voltage Pulse Quench the Fluorescence of Calcein in both Solution and Electroporated Cells

Abstract

To study cell electroporation, entrance into or efflux out of the cell of fluorescent dyes is often studied. However, when a cell suspension is exposed to high-voltage electric pulses, various electrochemical reactions occur at each electrode-solution interface. One of these reactions is the oxidation of the metal ions of the electrode. As a result of this, metal ions are released from the anode into the solution.In this study, influence of iron ions released from the stainless-steel anode on the fluorescence of the fluorescent molecules was studied. Calcein, which is used in studying cell electroporation, as well as anticancer drugs porphyrin-sulphonate and adriamycin, which can be used in combined application of the electroporation and photodynamic tumor therapy, were used.Iron ions suppressed the intensity of fluorescence of adriamycin, porphyrin-sulphonate, and calcein in solution. In the presence of 2.5 mM Fe3+, fluorescence of porphyrin sulphonate was suppressed by 100%, and adriamycin - by 50%. When mouse hematoma MH-22A cells preloaded with calcein were treated by a square-wave electric pulse with the duration of 2 ms and the amplitude of 1.6 kV/cm, a decrease in the number of fluorescent cells was observed: after the incubation for 5 min only 50% of cells were still fluorescent. However, when the cell suspension was supplemented with 0.55 mM FeCl3, quick decrease in the number of fluorescent cells was observed. When 5 min had passed after an electric pulse (2 min after the addition of Fe3+ ions), the fraction of fluorescent cells was only 25%.Therefore, the iron ions released from the stainless-steel anode can quench the fluorescence of the molecules not only in the solution but inside electroporated cells as well.