Sensitivity and Validation of Porous Membrane Electrical Cell Substrate Impedance Spectroscopy (PM-ECIS) for...

Area of Science:

• Biomedical Engineering
• Cell Biology
• Materials Science

Background:

• Endothelial and epithelial barrier integrity are crucial for in vitro models like Transwell assays and organ-on-chip platforms.
• Trans-endothelial electrical resistance (TEER) is the standard method for measuring barrier resistance but has limitations, including invasiveness and inability to measure isolated monolayer resistance in complex models.
• Porous membrane electrical cell-substrate impedance sensing (PM-ECIS) offers a noninvasive approach using electrodes on permeable membranes to assess barrier function.

Purpose of the Study:

• To advance the design and utility of PM-ECIS by investigating the impact of working electrode size on sensitivity and correlation with TEER.
• To optimize electrode selection for specific applications in cell barrier integrity measurements.
• To support PM-ECIS as a viable alternative to TEER for real-time assessment of cells on porous membranes.

Main Methods:

• Fabrication of gold electrodes with varying diameters (250, 500, 750 μm) on porous membrane inserts using hot embossing and UV lithography.
• Investigation of PM-ECIS sensitivity to resistance changes during endothelial barrier formation and cell spreading/proliferation at different frequencies (4 kHz and 40 kHz).
• Comparison of PM-ECIS measurements with TEER using sodium chloride solutions and endothelial monolayers.

Main Results:

• PM-ECIS sensitivity to resistance changes was inversely proportional to electrode size, with smaller electrodes (250 μm) showing the highest sensitivity (p < 0.001).
• Smaller electrodes were also more sensitive to impedance changes related to cell spreading and proliferation (p < 0.001).
• While all electrode sizes detected barrier disruption, PM-ECIS measurements correlated significantly with TEER for NaCl solutions (r > 0.9), and specifically with 750 μm electrodes for endothelial monolayers (r = 0.71, p = 0.058).

Conclusions:

• Electrode size is a critical design parameter for PM-ECIS, influencing sensitivity to barrier changes.
• Smaller electrodes are optimal for detecting subtle changes in barrier formation and cell behavior.
• PM-ECIS, particularly with optimized electrode selection, is a promising noninvasive alternative to TEER for real-time cell barrier assessment in diverse in vitro models.