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Content archived on 2024-06-16

Highly parallel cell culture in nanodrops, a new format for high content cell based toxicity screening on cell on chips

Final Report Summary - TOXDROP (Highly parallel cell culture in nanodrops, a new format for high content cell-based toxicity screening on cell on chips)

The ultimate aim of the TOXDROP project was to develop an innovative 'cell on chip' technology to screen cells for toxicity. For this reason it sought to provide an approach for automated high-throughput, high-content screening in toxicity assays using a revolutionary format of cellular nanodrops formed on a glass slide.

Two separate approaches were tested in order to facilitate toxicity analysis within these cellular nanodrops: On the one hand fluorescence microscopy was used to identify fluorescent proteins (fluorimetric assay). On the other hand two and three-dimensional time-of-flight secondary ion mass spectrometry analysis (TOF-SIMS) of cells was performed.

New software tools were developed for the analysis of the images provided by both approaches. Nanodrops were realised through conventional piezo-dispensing or through using a specific microfluidic device based on impedance detection of cells within the chip. Furthermore, fluorometric cell-based assays for global toxicity monitoring using the 'stress promoter technology' were developed. For this purpose, several cell lines of organs particularly sensitive to toxic chemicals were generated such as liver cells, lung bronchial epithelium cells, keratynocytes and differentiated hepatocytes. Especially, hepatocyte cells displayed active bio-transformation behaviour.

In order to perform the fluorometric assay, the dispensing conditions were set for the engineered cell lines and a 'cell on chip' device was used to analyse 400 independent nanodrops. By using ToF-SIMS it was possible to detect the three-dimensional (3D) distribution of molecules from both the cell membrane and the cell nucleus without any labelling. In addition, a xenobiotic fluorophor could be detected by fluorescence microscopy as well as by TOF-SIMS imaging, thus validating the technique. Cellular analysis and multiplex phenotypic characterisation was performed using IMSTAR Pathfinder leading technologies for high-content screening (HCS) analysis. By using this patented technology an automated and quick detection of cell borders, as well as fluorimetric response of cells was possible.

Thus, the enormous amount of data obtained by both, the fluorometric, as well as the mass spectrometric approach developed within the project could be evaluated. Given the large amount of information to process, a shared database was initiated to facilitate data storage and implementation of statistical tools.