Three-Chambered Array-Based Impedance Assay: A Real-Time Analysis Technique to Assess Invasive Potential of Cancer Cells by Measuring Electrical Impedance
Published: April 30, 2023
Abstract
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 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.
Protocol
1. New chamber design (Figure 1) Open a new dual-chamber cell analyzer plate. Set aside the top chamber with electrodes. Using a milling machine, shave off 2 mm of the U-shaped bottom wells of the cell analyzer plate. Attach a 2 cm x 7 cm polyethersulfone (PES) membrane with 0.2 μm 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. Using a milling machine, cut out two longitudina…
Representative Results
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…
Declarações
The authors have nothing to disclose.
Materials
| 0.05% Trypsin-EDTA | Thermofisher | 25300-054 | |
| Adhesive | Norland Optical Adhesive | NOA63 | |
| Bovine serum albumin (BSA) | Sigma | A9418 | |
| Cell lifter | Sarstedt | 83.1832 | |
| Cholera Toxin from Vibrio cholerae | Thermofisher | 12585-014 | |
| CIM-plate | Agilent 5665817001 | Cell analyzer plate | |
| Collagenase from Clostridium histolyticum | Sigma | C0130 | |
| DMEM | Thermofisher | 11995-065 | |
| DMEM-F12 | Thermofisher | 11875-093 | |
| Fetal Bovine Serum (FBS), Heat Inactivated | Omega Scientific | FB-12 | |
| HEPES | Thermofisher | 15630106 | |
| Horse serum (HS) | Gibco | 16050-122 | |
| Human EGF | Peprotech | AF-100-15 | |
| Hydrocortisone | Sigma | H4001 | |
| Insulin Transferrin Selenium Ethanolamine (ITSX) (100x) | Thermofisher | 51500056 | |
| Insulin, Human Recombinant, Zinc Solution | Sigma | C8052 | |
| J2 Fibroblasts | Stemcell (RRID:CVCL_W667) | 100-0353 | |
| LymphoPrep | Stemcell | 7851 | Density gradient medium for the isolation of mononuclear cells |
| Matrigel | Corning | 354230 | Basement membrane matrix |
| MCFDCIS.com cells ( DCIS) | RRID:CVCL_5552 | ||
| MDA-MB-231 cells | RRID:CVCL_0062 | ||
| Milling machine | Bridgeport Series 1 Vertical | ||
| Phosphate-buffered saline (1x) | Thermofisher | 10010049 | |
| Polyethersulfone (PES) membrane | Sterlitech | PCTF029030 | |
| RBC lysis solution | Stemcell | 7800 | |
| RTCA DP analyzer | Agilent | 3X16 | Dual purpose cell analyzer |
| Trypsin | Sigma | T4799 |
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Three-Chambered Array-Based Impedance Assay: A Real-Time Analysis Technique to Assess Invasive Potential of Cancer Cells by Measuring Electrical Impedance. J. Vis. Exp. (Pending Publication), e21156, doi: (2023).