Real-time Imaging and Tuning Subcellular Structures and Membrane Transport Kinetics of Single Live Cells at Nanosecond Regime

摘要

We developed an electric-field exposure microchannel system with 230-nanometer thin-layer gold electrodes, and interfaced it with a single living cell imaging station and a 10-nanosecond-electric-pulse (10nsEP) generator. This design allows us to image intracellular molecules and structures, membrane transport and viability of single leukemic cells (HL60) while the cells are exposed to 10nsEPs of 0–179 kV/cm, permitting the study of subcellular responses at nanosecond regime. The electrodes confine a thin-layer section of the cells exposed to 10nsEPs, offering unprecedented high spatial resolution (230-nm at z-direction of E and imaging plane) for imaging intracellular molecules of single cells affected by 10nsEPs. We found that nucleic acids, membrane transport rates and viability of single cells depend on the number and electric-field-strength (E) of 10nsEPs, showing the cumulative effect of 10nsEPs on intracellular molecules and structures and suggesting the possibility of tuning them one-pulse-at-a-time. Using lower E (51 kV/cm) of 10nsEPs, we could manipulate nucleic acids of single living cells without disrupting their cellular membrane and viability. As E increases to 80, 124 and 179 kV/cm, membrane integrity and viability of cells exhibit higher dependence on the number of 10nsEPs in a non-linear fashion, showing that critical E and pulse number are needed to surmount cellular transport barriers and membrane integrity.