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1. New chamber design (Figure 1)
<ol>
	<li>Open a new dual-chamber cell analyzer plate. Set aside the top chamber with electrodes.</li>
	<li>Using a milling machine, shave off 2 mm of the U-shaped bottom wells of the cell analyzer plate.</li>
	<li>Attach a 2 cm x 7 cm polyethersulfone (PES) membrane with 0.2 &#956;m pore size to the bottom of the shaved wells using UV-curated adhesive. Allow 30 min curation time to ensure the glue is completely cured and inert.</li>
	<li>Using a milling machine, cut out two longitudina...

three-chambered array-based impedance assay: a real-time analysis technique to assess invasive potential of cancer cells by measuring electrical impedance

Source: Sharif, G. M. et al., <a href="https://www.jove.com/v/63512">Real-Time Detection and Capture of Invasive Cell Subpopulations from Co-Cultures.</a> J. Vis. Exp. (2022).
This video describes an invasion assay using a three-chambered array based on electrical impedance. This technique measures the invasive potential of cancer cells under the influence of soluble factors secreted by the resident stromal cells in tumor microenvironments.

<img alt="Figure 1" src="/files/ftp_upload/21156/21156_Figure1.jpg" /> 
Figure 1: Images of the array chambers and modifications.
(A) The three chambers used to build the array. No modification was made on the top chamber harboring the electrodes. (B) From the middle chamber wells, a height of 2 mm has been shaved off and a membrane attached to the open bottom; longitudinal slits (1.5 mm x 5.6 mm) were add...

Three-Chambered Array-Based Impedance Assay: A Real-Time Analysis Technique to Assess Invasive Potential of Cancer Cells by Measuring Electrical Impedance

Source: Sharif, G. M. et al., Real-Time Detection and Capture of Invasive Cell Subpopulations from Co-Cultures. J. Vis. Exp. (2022).
This video describes ...

Cancer-associated stromal cells secrete chemical factors that can trigger the metastatic tumor cells residing on the tissue basement membrane. Consequently, the cancer cells invade the basement membrane and migrate toward the tissue&#39;s extracellular space.
To measure the invasive capacity of cancer cells in vitro, begin with a multiwell plate with multiple three-chambered arrays. In each array, the bottom chamber has an adherent culture of stromal cells. The top chamber with fused impedance-measuring microelectrodes - miniature electrodes that measure a cell&#39;s electrical activity - is overlaid by cancer cells, which adhere to the extracellular matrix-coated microporous membrane present above the electrodes.
During incubation on an impedance-based analyzer, the stromal cells release chemotactic factors into the medium, which move toward the cancer cells. These factors move through the semi-permeable membrane present at the bottom side of the middle chamber.
The stromal cell-secreted factors signal the adherent cancer cells to transform into invasive phenotypes. In response, the invasive cells migrate through the microporous membrane reaching the interdigitated electrodes on the opposite side of the membrane.
Over time, as more cancer cells stick to the electrodes, they cause cellular impedance - hindrance to the flow of current by cells. The measurement of cellular impedance is indicative of the invasive potential of cancer cells.

three-chambered-array-based-impedance-assay-real-time-analysis

Research

JoVE Encyclopedia of Experiments

Biological Techniques

Assay Techniques

Cell-based assays

This video describes an invasion assay using a three-chambered array based on electrical impedance. This technique measures the invasive potential of cancer cells under the influence of soluble factors secreted by the resident stromal cells in tumor microenvironments.

Overview

Protocol

<img title="figure-representative results-58" alt="figure-representative results-58" src="/files/ftp_upload/21156/21156_Figure1.jpg" /> 
Figure 1: Images of the array chambers and modifications.
(A) The three chambers used to build the array. No modification was made on the top chamber harboring the electrodes. (B) From the middle chamber wells, a height of 2 mm has been shaved off and a membrane attached to the open bottom; longitudinal slits (1.5 mm x 5.6 mm) were add...

Three-Chambered Array-Based Impedance Assay: A Real-Time Analysis Technique to Assess Invasive Potential of Cancer Cells by Measuring Electrical Impedance

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Overview

Protocol

Representative Results

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