| Abstract : |
Cell culture models are being used increasingly in the biotechnological field to study various biological and metabolic processes that happen in cells, tissues, organs and organisms as a whole.Here,cell populations are researched in vitroand isolated from surrounding tissues and their influences. Recently, these cell cultures have evolved from classic one-layer 2D models to more complex 3D models, in which several cell layers or even cell types are allowed to interact with each other. This to better replicate the physiology of tissues and organisms. In order to work with these complex cultures there is need for an accurate and trustworthy measurement technique that gives information about the cell number, viability, confluence and distribution of the cells within the culture. This measurement needs to happen in real-time and be non-destructive tothe sample so that the 3D cell culture can be monitored throughout its growth cycle. A promising upcoming technique is impedance spectroscopy, in which the culture is modelled asan electrical circuit. This technique is commonly used in 2D cell cultures buthas yet to be successfully implemented on3D cell culture models. Thisdue to the added complexity of their setup and the proper integration of the measurement electrodes within the culture. In this study the reproducibility and variability of impedancemeasurements performed on 2Dand2.5D was tested withthe electrodes placed directly beneath the culture. The cultures were also imaged to investigate the location of the cells and their structure relative to the electrodes. While this study was able tosuccessfullycompare 2D and 2.5D cell cultures in terms of variability, reproducibility and validation using brightfield microscopy images, some of the more complex experiments hadto be cancelled due to COVID-19. In regard to 2D cell cultures it was found that, while impedance spectroscopy is able to give a good representation of the observed cells in regards to dispersion and confluence of the cells, it is too sensitive to the natural variability in cells and cellcultures. Thismakes reproducibility andvalidation of the resultsdifficult due to the natural cell cycleand the cell location relative to the electrode surface. These challenges that already present themselves in 2D cell culturesmake the difficulty in using this measurement technique with 3Dcultures all the more apparent. The main challenge this paper brings forward is how thenatural occurring variability in cell cultures needs to be consideredwhen using the highly sensitive impedance spectroscopy. Future research should focus on quantifying this variability. This knowledge will likely prove useful in interpreting impedance data, which will helpwhen transferring these experiments from 2D to a more complex 3D culturemodel. If these challenges are overcome,the usage of impedance-based measurement systems will provide real-time, non-invasive and quantitative tracking of 3D cell culture systems.Keywords: Impedance spectroscopy, Brightfieldmicroscopy, 2D cell cultures, 2.5D cell cultures, 3D cell cultures. |
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