Toxicology with ECIS


The ECIS Method

For both economic and humane considerations there has been growing interest in alternatives to the use of animals in toxicity testing of chemical agents. Tissue culture has the potential to replace animal testing, but for such in vitro approaches to be successful, new and sensitive methods to detect cellular activities are required.

In 1992, we published an article suggesting the application of cell impedance measurement for this purpose [Giaever, I. and Keese, C.R., "Toxic? Cells Can Tell", Chemtech, 116-125 (Feb. 1992)]. Since that time, we have developed ECIS-based assays for this purpose and have acquired considerable data demonstrating the efficacy of this method to collect relevant toxicological data. The ECIS approach furnishes data that are by nature quantitative, and since the instrument is computer interfaced, very little technician labor is required to acquire large amounts of information. This sort of precision and cost effectiveness are important attributes of ECIS.

In 1998 dose response curves using ECIS were published [Keese, C., Karra, N., Dillon, B., Goldberg, A., Giaever, I., "Cell-Substratum Interactions as a Predictor of Cytotoxicity." In Vitro & Molecular Toxicology 11 (2), (1998)]. In this study cells in vitro were exposed to different levels of potential toxicants and their response monitored. Specifically, established confluent monolayers (WI-38/VA13 fibroblastic and MDCK epithelial cells) were exposed to varying concentrations of four detergents (Tween 20, benzalkonium chloride, Triton X100 and sodium lauryl sulfate). ECIS measurements were used to follow subsequent changes of the overall impedance of the cell monolayer and of cell micromotions detected as impedance fluctuations. Analysis of these measurements correctly ranked the detergents according to their established in vivo toxicity. Of interest in this work was a dramatic increase of impedance fluctuations sometimes recorded from cells upon exposure to the toxicants. This occurred when detergent concentrations were below those showing a decline in overall impedance; this hormesis effect was particularly evident in the MDCK cells. A portion of these published results is shown below.

The curves above show the time course change in the resistance (normalized to its value at time zero) upon exposure of confluent cells layers to detergent (type and concentrations indicated). These data exemplify the types of responses we have observed. At high concentrations of detergent, a rapid drop in the resistive portion of the impedance is observed that begins near the time of detergent addition. At intermediate concentrations there is a delay in response followed by a slower decline in the resistance Of special interest is the hormesis effect evident in the increase in impedance when MDCK cells were exposed to 500 micrograms per ml of Tween 20.
Dose response curves of WI38/VA13 cells exposed to four different types of detergents. Healthy confluent cell layers were exposed to detergents for 20 hours. At the end of this time, the in-phase voltage (related to the resistive portion of the impedance) was measured each second until 1024 points were acquired. The average value of this quantity is shown as a percentage of that measured from control cultures. Also indicated in the average statistical variance in the fluctuations of the in-phase voltage measured in 32 second increments compared as a percentage of that measured from controls.

Toxicology
Related ECIS Publications

Electric cell-substrate impedance sensing (ECIS) based real-time measurement of titer dependent cytotoxicity induced by adenoviral vectors in an IPI-2I cell culture model. J Muller, C Thirion, and MW Pfaffl. Biosens Bioelectron. 2010.

Development of an electrode cell impedance method to measure osteoblast cell activity in magnesium-conditioned media. Y Yun, Z Dong, Z Tan, and MJ Schulz. Anal Bioanal Chem. 2010. Volume: 396   Issue: 8 Pages: 3009-3015.

Theresa M. Curtis, Mark W. Widder, Linda M. Brennan, Steven J. Schwager, William H. van der Schalie, Julien Fey and Noe Salazar,  “A portable cell-based impedance sensor for toxicity testing of drinking water.” Lab Chip, 2009, DOI: 10.1039/b901314h.

Theresa M. Curtis, Joel Tabb, Lori Romeo, Steven J. Schwager, Mark W. Widder and William H. van der Schaliec, Improved cell sensitivity and longevity in a rapid impedance-based toxicity sensor.” Journal of Applied Toxicology (January 2009).

Campbell, C.E., Motzfeldt-Laane, M., Haugarvoll, E., and Giaever, I., "Monitoring Viral Induced Cell Death using Electric Cell-Substrate Impedance Sensing." Biosensors and Bioelectronics: 23(4): 536-542 (2007).

van der Schalie, W.H,. James, R.R., Gargan, T.P. 2nd, "Selection of a battery of rapid toxicity sensors for drinking water evaluation." Biosens Bioelectron, (2006).

Chanana, M., Gliozzi, A., Diaspro, A., Chodnevskaja, I., Huewel, S., Moskalenko, V., Ulrichs, K., Galla, H.J., Krol, S., "Interaction of polyelectrolytes and their composites with living cells." Nano Lett. 5(12):2605-12 (2005).

Xiao, C., Luong, J.HT., "The Assessment of Cytotoxicity by Emerging Impedance Spectroscopy." Toxicology and Applied Pharmacology, (2004).

Xiao, C., Luong, J., "On-Line Monitoring of Cell Growth and Cytotoxicity Using Electric cell-substrate Impedance Sensing (ECIS)." ,Biotechnal Prog, 19, 1000-2005 (2003)

Xiao,C., Lachance, B., Sunahara, G., Luong, J., "Assessment of Cytotoxicity Using Electric Cell-Substrate Impedance Sensing: Concentration and Time Response Function Approach.", Anal. Chem., 74(22), 5748-5753 (2002)

Noiri, E., Nakao, A., Uchida, K., Tsukahara, H., Ohno, M., Fujita, T., Brodsky, S., Goligorsky, M., "Oxidative and nitrosative stress in acute renal ischemia", Am J Physiol Renal Physiol 281: F948-F957 (2001)

Keese, C., Karra, N., Dillon, B., Goldberg, A., Giaever, I., "Cell-Substratum Interactions as a Predictor of Cytotoxixity." In Vitro & Molecular Toxicology 11 (2), (1998)

Giaever, I. and Keese, C.R., "Toxic? Cells Can Tell", Chemtech, 116-125 (Feb. 1992).