Department of Electrical and Computer Engineering, University of Manitoba, Winnipeg, Canada.
Department of Microbiology, University of Manitoba, Winnipeg, Canada.
Anal Chim Acta. 2019 Jun 20;1059:59-67. doi: 10.1016/j.aca.2019.01.046. Epub 2019 Feb 3.
The dielectric properties of cells are directly related to their morphological and physiological properties and can be used to monitor their status when exposed to stress conditions. In this work, dual-frequency dielectrophoresis (DEP) cytometry was employed to measure changes in the membrane capacitance and cytoplasm conductivity of single Chinese hamster ovary (CHO) cells during the progression of starvation-induced apoptosis. Our dual-frequency DEP cytometer enables simultaneous measurement of multiple dielectric properties of single cells and identification of their state (viable or apoptotic) within a heterogeneous sample. We employed one frequency to determine each cell's viability state and the other frequency to characterize the change in membrane capacitance or cytoplasm conductivity. Cells were starved by incubation in a medium lacking glucose and glutamine and monitored every 12 h over a 64 h period. Our results showed a subpopulation of early apoptotic cells emerged after 40 h in the starvation medium, which rapidly increased during the next 12 h. After 52 h, a complete transition from viable to apoptotic state was observed. Analyzing the subpopulation of viable cells over the first 52 h showed that the membrane capacitance gradually declined from an initial value of 2.0 to 1.2 μF/cm, and was 0.9 μF/cm for apoptotic cells. The cytoplasm conductivity of viable cells initially remained constant and then declined from 0.40 to 0.27 S/m after 40 h, coinciding with onset of apoptotic processes. A dramatic decrease in cytoplasm conductivity from 0.27 to 0.07 S/m was observed after 52 h, corresponding to apoptotic cells. As membrane capacitance is related to membrane morphology and cytoplasm conductivity is related to intracellular ion concentrations, the results indicate that during controlled starvation the cell membrane smooths gradually whereas intracellular ion concentrations are initially maintained near homeostatic levels until a later dramatic decline occurs.
细胞的介电特性与其形态和生理特性直接相关，可用于监测细胞在应激条件下的状态。在这项工作中，采用双频介电泳（DEP）细胞术测量了饥饿诱导的细胞凋亡过程中单个中国仓鼠卵巢（CHO）细胞的膜电容和细胞质电导率的变化。我们的双频 DEP 细胞仪能够同时测量单个细胞的多种介电特性，并在异质样品中识别其状态（存活或凋亡）。我们使用一个频率来确定每个细胞的存活状态，另一个频率来描述膜电容或细胞质电导率的变化。通过在缺乏葡萄糖和谷氨酰胺的培养基中孵育来使细胞饥饿，并在 64 小时的过程中每 12 小时监测一次。我们的结果表明，在饥饿培养基中 40 小时后出现了一小部分早期凋亡细胞，在接下来的 12 小时内迅速增加。在 52 小时后，观察到从存活状态到凋亡状态的完全转变。分析前 52 小时内存活细胞的亚群表明，膜电容从初始值 2.0 逐渐下降到 1.2 μF/cm，而凋亡细胞的膜电容为 0.9 μF/cm。存活细胞的细胞质电导率最初保持不变，然后在 40 小时后从 0.40 下降到 0.27 S/m，与凋亡过程的开始相吻合。在 52 小时后，细胞质电导率从 0.27 急剧下降到 0.07 S/m，对应于凋亡细胞。由于膜电容与膜形态有关，细胞质电导率与细胞内离子浓度有关，因此结果表明，在受控饥饿过程中，细胞膜逐渐变平，而细胞内离子浓度最初保持在接近体内平衡的水平，直到后来发生急剧下降。
