Anionic Conjugated Polyelectrolytes for FRET‐based Imaging of Cellular Membrane Potential

We report a Forster resonance energy transfer (FRET)-based imaging ensemble for the visualization of membrane potential in living cells. A water-soluble poly(fluorene-co-phenylene) conjugated polyelectrolyte (FsPFc10) serves as a FRET donor to a voltage-sensitive dye acceptor (FluoVolt(TM) ). We observe FRET between FsPFc10 and FluoVolt(TM) , where the enhancement in FRET-sensitized emission from FluoVolt(TM) is measured at various donor/acceptor ratios. At a donor/acceptor ratio of 1, the excitation of FluoVolt(TM) in a FRET configuration results in a 3-fold enhancement in its fluorescence emission (compared to when it is excited directly). FsPFc10 efficiently labels the plasma membrane of HEK 293T/17 cells and remains resident with minimal cellular internalization for ~1.5 h. The successful plasma membrane-associated co-labeling of the cells with the FsPFc10-FluoVolt(TM) donor-acceptor pair is confirmed by dual channel confocal imaging. Importantly, cells labeled with FsPFc10 show excellent cellular viability with no adverse effect on cell membrane depolarization. During depolarization of membrane potential, HEK 293T/17 cells labeled with the donor-acceptor FRET pair exhibit a greater fluorescence response in FluoVolt(TM) emission relative to when FluoVolt(TM) is used as the sole imaging probe. These results demonstrate the conjugated polyelectrolyte to be a new class of membrane-labeling fluorophore for use in voltage sensing schemes.