Miniaturized and parallelized video microscopy for 96-well microplates – CellScan

The parallelised holographic microscope includes a multi-sensors image acquisition system, an illumination system and a mechanical support for the 96-well microtiter plate which is connected to XY motorised positioners. The components were assembled in an internally-designed housing whose materials and fabrication techniques are compatible with serial production. The images are automatically recorded according to the time rate and duration specified by the user. Software was programmed to visualise and process the 96 image series supplied by the microscope. Furthermore, dedicated software was programmed for rapid and user-friendly analysis of the results of a screen, e.g., motility measurements using wound healing assays. The microscope and software environment allowed identifying and validating several hits during the project.

The microscope was successfully used in high-content screens. The imaging system is compatible with every format of microtiter plates and every standard culture substrate. The printed circuit boards are very stable and support the massive acquisition of 96 image series. The automatic segmentation of objects on the images was successfully compared to a campaign of manual segmentations performed by several users. Internal users warmly welcomed software components for image visualization and graphical analysis of quantitative data.

The video microscope prototype and software are fully operational. Our first screens have uncovered some hit molecules which will be specifically investigated in the near future. These specific results validate the general approach of our video microscopy system for the screening application. The preproduction phase of the holographic video microscope will start soon.

1) V. Haguet et al., Proc. 17th MicroTAS Conference, pp. 1740-1742, Freiburg, Germany, October 27-31, 2013.
2) I. Ghorbel et al., Proc. 37th International Meeting of the German Society for Cell Biology (DGZ), p. 71, Regensburg, Germany, March 18-21, 2014.

Standard imaging of chemically fixed cells offers valuable information on cellular structure and phenotype to biology academics and the pharmaceutical industry. However, it provides limited means to disclose kinetic data such as transient or rare cellular events. There is currently a growing demand for monitoring the evolution of cell cultures to identify specific cellular signatures, e.g. recording drug effects on cell viability, proliferation, morphology or motility. Video microscopy instruments and recent imaging devices in an incubator environment can provide time-lapse imaging of living cells. Nevertheless, these solutions are costly (in the 50-190 k€ range) and/or have no or low-speed parallelization capabilities. They frequently supply restricted fields of view, medium resolution, and only monofluorescence abilities due to cost reasons. In this 18-month project, we aim at applying our previously developed contact-mode cell imaging technology to tackle these issues. Our disruptive imaging architecture with no costly optical or contrast-enhancement components can generate time-lapse bright-field images of cells in microplates inside an incubator. The laboratories BGE and LISA of the CEA in Grenoble will incorporate new added-valued capabilities to the parallelized cell imaging device based on a centimeter-travel micropositioning system, thin-film optical coatings and image processing software: (i) visualizing the whole area of microplate wells so as to exhaustively screen the cell populations; (ii) imaging fluorescent cells using interference filters micropatterned on the image sensors; (iii) massive automated data collection onto the cells by high-throughput image segmentation and analysis; (iv) subcellular imaging based on holographic image reconstruction software and specific diffraction-enhancement labelling. With the help of the CEA Technology Transfer Office, the valorization objectives will be to extend patent protection to the device architecture and the operating algorithms and methods, and organize technology transfer through licensing to a private company in the imagery field, possibly to a spin-off company if this approach is financially justified.