In vitro Intestinal Epithelial Wound-healing Assays Using Electric Cell-Substrate Impedance Sensing Instrument
In vitro Intestinal Epithelial Wound-healing Assays Using Electric Cell-Substrate Impedance Sensing Instrument
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DOI：
10.21769/bioprotoc.3351
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发表时间：
2019-09-05
期刊：
影响因子：
0.8
通讯作者：
Viennois, Emilie
中科院分区：
文献类型：
--
作者：
Merlin-Zhang, Olivier;Sung, Junsik;Viennois, Emilie
在这里，我们描述了一种体外上皮伤口愈合测定使用电细胞基质阻抗传感（ECIS）技术。ECIS技术是一种真实的时间细胞生长测定法，其基于小的（250 μ m直径）活性金电极，其电阻被连续测量。当肠上皮细胞在金电极上达到汇合时，电阻达到平台。对于伤口愈合测定，使汇合的肠上皮单层经受40 kHz频率、1，400 μ A振幅和30秒持续时间的电流。这杀死了小的活性金电极周围的细胞，导致分离并产生伤口，伤口被周围没有受到电流脉冲的细胞愈合。然后通过在伤口后持续约30小时的电阻测量来评估伤口愈合。细胞损伤和后续愈合过程的测量都是在计算机控制下进行的，每30秒进行一次在线测量并存储数据。ECIS技术可用于研究粘膜愈合受损的根本原因，并测试药物在粘膜愈合中的功效。
Here, we describe an in vitro epithelial wound-healing assay using Electric Cell-Substrate Impedance Sensing (ECIS) technology. The ECIS technology is a real time cell growth assay based on a small (250 mu m diameter) active gold electrode which resistance is measured continuously. When intestinal epithelial cells reach confluency on the gold electrode, resistances reach a plateau. For the wound-healing assays, confluent intestinal epithelial monolayers are subjected to a current of 40 kHz frequency, 1,400 mu A amplitude, and 30-second duration. This kills the cells around the small active gold electrode, causing detachment and generating a wound that is healed by surrounding cells that have not been submitted to the current pulse. Wound healing is then assessed by continuous resistance measurements for approximately 30 h after wound. Both cell wounding and measurements of the subsequent healing process are carried out under computer control that takes online measurements each 30 s and stores the data. ECIS technology can be used to study the underlying causes for impaired mucosal healing and to test the efficacy of drugs in mucosal healing.