The membrane potential of a cell is defined as the voltage inside the cell relative to the extracellular solution. Preexisting methods to measure cell membrane potential are more invasive and harmful to cells compared to the process of fluorescence resonant energy transfer (FRET). FRET utilizes a donor-acceptor energy transfer molecular pair in order to reflect changes in membrane potential. My research examined the use of the energy-transfer pair DiOC18(3) (a fluorescent tracer dye) and DPA (a hydrophobic anion) as an optical voltage sensor. Using DiOC18(3) and DPA in REF52 cells, we recorded the initial fluorescence quenching of FRET with fluorescence microscopy; however, no decrease in fluorescence quenching was detectable when the cell membrane potential was altered with a high concentration potassium solution (60 mM KCl). These results indicate that the REF52 cells were successfully labeled with DiOC18(3) and that DPA could act as an energy acceptor and quench DiOC18(3) fluorescence through FRET. Although we know from physical principles that the efficiency of FRET should be altered when a high K+ solution is applied, no change was observed. Therefore, we concluded that the change in fluorescence upon depolarization was simply too small to be reliably detected. These findings validate the importance of alternative voltage sensing methods and provide a possible model for next-generation energy-transfer pairs. In this manner, cell membrane voltage differences can be measured without harming cells.
See-yin So ’10 Wichita, KS
Majors: Biochemistry and Molecular Biology, Ethnic Studies
Sponsor: Craig Tepper