Method for Extracellular Electrochemical Impedance Spectroscopy on Epithelial Cell Monolayers

Area of Science:

• Epithelial physiology
• Electrophysiology
• Biophysics

Background:

• Epithelial tissues are crucial barriers, and transepithelial resistance (TER) is a common measure of their integrity.
• Conventional TER measurements do not fully capture the complex electrical behavior or membrane-specific properties of epithelia.

Purpose of the Study:

• To present a protocol for galvanostatic electrochemical impedance spectroscopy (EIS) to measure epithelial electrical properties.
• To enable the measurement of transepithelial capacitance (TEC) and a new metric, the membrane ratio, alongside TER.

Main Methods:

• Utilized galvanostatic EIS with a 4 μA amplitude signal across a 2 Hz to 50 kHz frequency range.
• Employed commercially available cell culture inserts and chambers for measurements.
• Developed a fitting technique with quantitative error estimation, including provided MATLAB code.

Main Results:

• Demonstrated repeatable measurements with mean absolute error below 10 Ω.
• Obtained distinct TER, TEC, and membrane ratio values for retinal pigment and bronchiolar epithelial samples.
• TER values ranged from 500-1034 Ω·cm², TEC from 1.07-4.10 μF/cm², and membrane ratios from 1.9-22.

Conclusions:

• Galvanostatic EIS provides a rapid (under 1 min) and detailed characterization of epithelial electrical properties.
• This method offers a more complete assessment of epithelial barrier function than TER alone.
• The protocol is adaptable for various epithelial cell types and allows for repeated measurements on the same sample.