Impedance-Based Bioassay for Characterization of Single Malignant Melanoma Cancer Cells using Cmos-Mea Systems

Abstract

Malignant Melanoma (MM) is the most aggressive type of skin-cancer. Current diagnostic tools for the detection of malignancies of the skin (MM cancer) include histological, optical, ultrasound, and impedance-based techniques. The inadequacies of the first three practices are overwhelmed by the Electrical Impedance Spectroscopy (EIS) method. EIS overcomes reported spatiotemporal tradeoffs as a label-free and optics-free analytical method. Yet, MM’s enhanced heterogeneity and metastatic potential still results in misdiagnosis, or late diagnosis leading to stages characterized by high mortality rates. Important biological information and processing ability on single-cell level is missing. Single-cell dynamics recorded with a high-throughput system, contain important biological information on the heterogeneous subpopulations which are responsible for the MM aggressiveness.This project aims to investigate experimentally the possibility and capabilities of such a bioassay development, create working protocols and generate a fundamental basis for analysis and interpretation of the big-data-sets which derive from Impedance monitoring from a high-throughput transducer.Experiments were performed, employing two diverse, human-derived, MM cancer cell-lines, and using a high-throughput HD-MEA system with a 1024-channel impedance readout unit developed at IMEC, in Belgium. The measurements were realized at 1kHz aiming to extract Rseal information. The main proposal presents an experimental protocol of mid-term and long-term experiments Temporal and spatial resolutions were enhanced (Control System Automation), allowing for implementation of an experimental set to test the assay’s capabilities and determine any necessary additions to make the assay more robust for research (i.e. Z-Map, templates and scripts for OriginLab and Matlab, statistical methods for validation of findings on the big-data sets, optimizations in the experimental process, etc).Experimental results proved the optics-free specification of the assay with the utilization of an Impedance colormap, which was validated by comparing it to confocal images after measurements. Electrode-size and normalization techniques were deemed crucial factors that affected the variance of the results for the largest and smallest electrode sizes. Confluence level (70% or 10%) of the cell on the chip was an added biasing factor. IM measurements at 1kHz for two MM cell-lines (MM087 & MM029) showed a good possibility for classification (long-term experiments), while the biological findings on the heterogeneity information (mid-term and long-term experiments) cannot be considered conclusive until a full-scale analysis is conducted on the data. A proposal for a new type of analysis on the Impedance data is presented; it is an example-based approach to extract the classification and heterogeneity information of normalized IM measurements, by obtaining and analyzing the trends of impedance variance over time, per electrode.The bioassay proposal that was developed and tested in this project shows potential for both classification and heterogeneity studies for MM cancer. Further experimentation and development of validation techniques is required.