Max Planck Institute for Dynamics and Self-Organisation, Göttingen, Germany.
Methods Mol Biol. 2024;2828:119-145. doi: 10.1007/978-1-0716-4023-4_11.
The electric cell-substrate impedance sensing (ECIS) is a well-established technique that allows for the real-time monitoring of cell cultures growing on gold-electrodes embedded in culture dishes. Its foundation lays on the insulating effect that cells present against the free-flow of electrons, as these passive electrical properties generate a characteristic complex impedance spectrum when a small-amplitude, non-invasive alternating current (AC) is provided through the electrodes, the living cells, and the culture media in the culture ware. In addition, it possesses the ability to create a wound that is highly confined to the electrode area by simply increasing the amplitude of the AC current in dependence of the pre-resistor strength for a defined pulse duration and at a specific frequency. Therefore, it represents a controlled and reproducible tool to carry out in vitro wound healing experiments. Accordingly, in this methods protocol, the use of the ECIS will be described in the context of the wound healing research: cardiac 3T3 fibroblasts will be wounded and their recovery dynamics analyzed based on the typical methodologies applied to the processing of ECIS data. In addition, cellular micromotions will be evaluated. Finally, fluorescence immunostaining of ECIS samples will be described in order to showcase the potential of the ECIS in combination with other well-established techniques to add further knowledge depth to the understanding of the complex wound healing dynamics.
细胞-基质阻抗传感（ECIS）是一种成熟的技术，可实时监测嵌入培养皿中的金电极上生长的细胞培养物。其基础是细胞对电子自由流动的绝缘效应，因为这些被动电特性在通过电极、活细胞和培养容器中的培养基提供小幅度、非侵入性交流电（AC）时会产生特征性的复阻抗谱。此外，它还能够通过简单地增加 AC 电流的幅度来产生高度局限于电极区域的伤口，这取决于预电阻器的强度、定义的脉冲持续时间和特定的频率。因此，它代表了一种可控制和可重复的工具，可用于进行体外伤口愈合实验。因此，在本方法方案中，将在伤口愈合研究的背景下描述 ECIS 的使用：将心脏 3T3 成纤维细胞受伤，并根据应用于 ECIS 数据处理的典型方法分析其恢复动态。此外，还将评估细胞微运动。最后，将描述 ECIS 样品的荧光免疫染色，以展示 ECIS 与其他成熟技术相结合的潜力，为理解复杂的伤口愈合动态增加更多的知识深度。
