Department of Chemistry and Biochemistry, Old Dominion University, Norfolk, Virginia 23529, USA.
J Phys Chem B. 2009 Oct 29;113(43):14393-404. doi: 10.1021/jp9021739.
We developed an electric-field exposure microchannel system with 230-nm thin-layer gold electrodes and interfaced it with a single living cell imaging station and a 10-ns electric-pulse (10 nsEP) 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 10 nsEPs of 0-179 kV/cm, permitting the study of subcellular responses within a nanosecond regime. The electrodes confine a thin-layer section of the cells exposed to 10 nsEPs, offering unprecedented high spatial resolution (230 nm in the z-direction of imaging plane and electric field) for imaging intracellular molecules of single cells affected by 10 nsEPs. We found that nucleic acids, membrane transport rates, and viability of single cells depend on the number and electric-field-strength (E) of 10 nsEPs, showing the cumulative effect of 10 nsEPs on intracellular molecules and structures and suggesting the possibility of tuning them one-pulse-at-a-time. Using a lower E (51 kV/cm) of 10 nsEPs, 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 10 nsEPs in a nonlinear fashion, showing that a critical E and pulse number are needed to surmount cellular transport barriers and membrane integrity.
我们开发了一个电场暴露微通道系统，该系统具有 230nm 厚的金电极，并与单个活细胞成像站和 10ns 电脉冲（10nsEP）发生器相连接。这种设计使我们能够在细胞暴露于 0-179kV/cm 的 10nsEP 时对细胞内分子和结构、膜转运以及单个白血病细胞（HL60）的活力进行成像，从而可以在纳秒范围内研究亚细胞反应。该电极将细胞的一个薄层暴露于 10nsEP 中，为受 10nsEP 影响的单个细胞内分子的成像提供了前所未有的高空间分辨率（成像平面和电场的 z 方向为 230nm）。我们发现，单个细胞的核酸、膜转运率和活力取决于 10nsEP 的数量和电场强度（E），这表明 10nsEP 对细胞内分子和结构的累积效应，并提示可以逐个脉冲地调整它们。使用较低的 10nsEP 电场（51kV/cm），我们可以在不破坏细胞膜和活力的情况下操纵单个活细胞的核酸。随着 E 增加到 80、124 和 179kV/cm，细胞的膜完整性和活力对 10nsEP 数量的依赖性呈现出非线性的更高程度，表明需要一个临界的 E 和脉冲数来克服细胞转运障碍和膜完整性。
