Name: Electrical Cell-Substrate Impedance Sensing (ECIS)
Uploaded: 2023-05-26T12:30:00
Description: Watch this Scientific Journal Video about Electrical Cell-Substrate Impedance Sensing ECIS at JoVE.com

electrical cell-substrate impedance sensing (ecis)

通过电电池-衬底传感评估 MOF 诱导的电池行为变化

Metal-organic frameworks (MOFs) are hybrids formed through the combination of metal ions and organic linkers1,2 in organic solvents. Due to the variety of such combinations, MOFs possess structural diversity3, tunable porosity, high thermal stability, and high surface areas4,5. Such characteristics make them attractive candidates in a variety of applications, from gas storage6,7 to catalysis8,9, and from contrast agents10,11 to drug delivery units12,13. However, the implementation of MOFs into such applications has raised concerns relative to their safety to both the users and the environment. Preliminary studies have shown, for instance, that cellular function and growth change upon the exposure of cells to metal ions or linkers used for MOF synthesis1,14,15. For instance, Tamames-Tabar et al. demonstrated that ZIF-8 MOF, a Zn-based MOF, was leading to more cellular changes in a human cervical cancer cell line (HeLa) and a mouse macrophage cell line (J774) relative to Zr-based and Fe-based MOFs. Such effects were&#160;presumably due to the metal component of ZIF-8 (i.e., Zn), which could potentially induce cell apoptosis upon framework disintegration and Zn ion release1. Similarly, Gandara-Loe et al. demonstrated that HKUST-1, a Cu-based MOF, caused the highest reduction in mouse retinoblastoma cell viability when used at concentrations of 10 &#181;g/mL or greater. This was presumably due to the Cu metal ion incorporated during the synthesis of this framework, which, once released, could induce oxidative stress in the exposed cells15.
Moreover, analysis showed that the exposure&#160;to MOFs with different physicochemical characteristics could lead to varying responses of exposed cells. For instance, Wagner et al. demonstrated that ZIF-8 and MIL-160 (an Al-based framework), used in the exposure of an immortalized human bronchial epithelial cell, led to cellular responses dependent on frameworks&#39; physicochemical properties, namely hydrophobicity, size, and structural characteristics16. Complementarily, Chen et al. demonstrated that a concentration of 160 &#181;g/mL MIL-100(Fe) exposed to human normal liver cells (HL-7702) caused the largest loss in cellular viability, presumably due to the metal component of this specific framework (i.e., Fe17).
While these studies categorize MOFs&#39; deleterious effects on cellular systems based on their physicochemical characteristics and exposure concentrations, thus raising potential concerns with framework implementation, especially in biomedical fields, most of these evaluations are based on single time point colorimetric assays. For instance, it was shown that when (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium (MTT) and water-soluble tetrazolium salt (WST-1) assays were used, these biochemical reagents could lead to false positives upon&#160;their&#160;interactions with the particles that the cells were also exposed to18. The tetrazolium salt and neutral red reagents were&#160;shown to possess a high adsorption or binding affinity onto the surfaces of the particles, resulting to agent signal interference19. Moreover, for other types of assays, such as flow cytometry, which was previously shown to be used for assessing changes in cells exposed to MOFs20,21, it was shown that major issues have to be circumvented if a viable analysis of particles&#39; deleterious effects is to be considered. In particular, detection ranges of the particles&#39; sizes, especially in mixed populations like the ones offered by MOFs or references of the particles used for calibration before cellular changes, have to be addressed22. It was also shown that the dye used during cell labeling for such cytometry assays could also interfere with the nanoparticles that the cells were exposed to23.
The goal of this study was to use a real-time, high-throughput evaluation assay to assess changes in cell behavior upon exposure to a select MOF. Real-time evaluations can help provide insights into time-dependent effects, as related to the windows of exposures16. Further, they provide information on changes in cell-substrate interactions, cell morphology, and cell-cell interactions, as well as how such changes depend on the physicochemical properties of the materials of interest and exposure times24,25 respectively.
To demonstrate the validity and applicability of the proposed approach, human bronchial epithelial (BEAS-2B) cells, ZIF-8 (a hydrophobic framework of zeolitic imidazolate16), and electric cell-substrate impedance sensing (ECIS) were used. BEAS-2B cells represent a model for lung exposure26 and have been previously used to evaluate changes upon the exposure of cells to nanoclays and their thermally degraded byproducts26,27,28,&#160;as well as assess the toxicity of nanomaterials, such as single-walled carbon nanotubes (SWCNTs)18. Furthermore, such cells have been used for more than 30 years as a model for pulmonary epithelial function29. ZIF-8 was chosen due to its wide implementation&#160;in catalysis30 and as contrast agents31 for bioimaging and drug delivery32, and thus for the extended potential for lung exposure during such applications. Lastly, ECIS, the noninvasive, real-time technique, was previously used to evaluate changes in cell adherence, proliferation, motility, and morphology16,26 as a result of a variety of interactions between analytes (both materials and drugs) and exposed cells in real-time16,18,28. ECIS uses an alternating current (AC) to measure the impedance of cells immobilized on gold electrodes, with the impedance changes giving insights into changes in resistance and capacitance at the&#160;cell-gold substrate interface, barrier function as induced by cell-cell interactions, and over-cell layer coverage of such gold electrodes33,34. Using ECIS allows quantitative measurements at a nanoscale resolution in a noninvasive, real-time manner26,34.
This study assesses and compares the simplicity and ease of evaluation of MOF-induced changes in cellular behavior in real-time with single-point assay evaluations. Such a study could be further extrapolated for evaluating cell profiles in response to exposure to other particles of interest, thus allowing for safe-by-design particle testing and subsequent helping with&#160;implementation. Moreover, this study could complement genetic and cellular assays that are single-point evaluations. This could lead to a more informed analysis of the deleterious effects of particles on the cellular population and could be used for screening such particles&#39; toxicity in a high-throughput manner16,35,36.

作者聚焦：评估人肺上皮细胞对金属有机框架反应的进展 

用于实时评估金属有机框架毒理学特征的电池-基底传感

My research aims to evaluate changes in human lung epithelial cells'behavior upon exposure to metal organic framework, hybrids formed by combining metal ions in organic linkers in organic solvents. This research demonstrated that the physical and chemical composition of metal organic frameworks, as well as their cellular exposure ratios, affects the behavior of human lung epithelial cells in a time and dose-dependent manner. This can be recorded in a high throughput in real time fashion.Our research helps to evaluate, understand the mechanisms of toxicity of metal organic frameworks upon exposure to cellular systems. The current method allows continuous cell monitoring in a real time and in high throughput manner. Additionally, this method does not rely on any external reagents, and thus it reduces any possible interference they could have with the frameworks used for exposure.Our laboratory will continue to focus on understanding and evaluating the mechanisms of toxicity associated with nanomaterials exposures, as well as using such information to synthesize new and safe systems that could be implemented in biomedical applications at minimum risks to the user or the environment.

Watch this Scientific Journal Video about Electrical Cell-Substrate Impedance Sensing ECIS at JoVE.com

Electrical Cell-Substrate Impedance Sensing (ECIS)  

电电池-基板阻抗检测 （ECIS）

首先，取一个 96 孔板，其中包含覆盖约 4 平方毫米面积的叉指连接电极。将板放在井站上，然后单击设置按钮。如果井连接正确，将显示绿色读数。任何未连接的井都将被标识为红色。向正在使用的每个孔中加入200微升培养基，然后将板放入电池基板阻抗传感或ECIS站中。要稳定孔并为实验做好准备，请单击设置，然后单击检查，然后单击稳定。40分钟后，从工作站上取下板并从每个孔中取出培养基。将B2B细胞以每毫升40， 000个细胞的密度接种到含有细胞的孔中，并将板放在培养箱中的ECIS站上。接下来，单击检查以确保工作站上的传感器读取所有电极。选择多频率时间以七个不同频率测量每个电极。单击开始并允许它运行 24 小时，以便细胞可以形成融合的单层。每口井的读数都以不同的颜色显示。大约24小时后，含有细胞的孔将在监视器上显示增加的电阻，表明细胞附着在电极上。按暂停可中断数据收集。例如，对于介质更改、试剂版本等。将ZIF8纳米颗粒暴露在空白孔和每孔体积为100微升的细胞孔中，将板放在工作站上，然后再次单击检查以确保传感器正在读取所有电极。单击“恢复”可在所选暂停后继续实验。数据收集后，单击完成将数据另存为电子表格文件。要获取 alpha 参数，请单击模型，然后在弹出的屏幕上，单击仅媒体井，然后选择只有介质的井。单击设置数据评估参考，然后选择与对照实验相关的参考。单击“查找”（Find） 以启动参照管接头。选择“设置”，然后选择“拟合 T 范围”以处理相对于所选引用的数据。若要评估单元格附件，请选择 alpha。最后，在系统完成分析后，单击保存 RBA 并将参数另存为电子表格文件。通过电池基板阻抗传感对暴露于ZIF8 MOFs时的电池行为进行分析。图像显示了细胞底物相互作用的变化，这些变化取决于暴露时间和使用的剂量。

Research

JoVE Journal

Bioengineering

Electrical Cell-Substrate Impedance Sensing (ECIS)

Electric Cell-Substrate Sensing for Real-Time Evaluation of Metal-Organic Framework Toxicological Profiles

Metal-organic frameworks (MOFs) are hybrids formed through the coordination of metal ions and organic linkers in organic solvents. The implementation of ...