| Popis: |
Research is continuously developing imaging methods to better understand the structure and function of biological systems. In this paper, we describe our work to develop lens-free microscopy as a novel mean to observe and quantify cells in 2D and 3D cell culture conditions. At first, we developed a lens-free video microscope based on multiple wavelength acquisitions to perform time-lapse 2D imaging of dense cell culture inside the incubator. We demonstrated that novel phase retrieval techniques enable imaging thin cell samples with high concentration (~15000 cells over a large field of view of 29.4 mm2). The experimental data can next be further analyzed with existing cell profiling and tracking algorithms. As an example, we showed that a 7 days acquisition of a culture of HeLa cells leads to a dataset featuring 2.106 cell point measurements and 104 cell cycle tracks. Recently, we extended our work to the video-microscopy of 3D organoids culture. We showed the capability of lens-free microscopy to perform 3D+time acquisitions of 3D organoids culture. To our knowledge, our technique is the only one able to reconstruct very large volumes of 3D cell culture (~5 mm3) by phase contrast imaging. This new mean of microscopy allowed us to observe a broad range of phenomena present in 3D environments, e.g. self-organizations, displacement of large clusters, merging and interconnection over long distances (>1 mm). In addition, this 3D microscope can capture the interactions of single cells and organoids with their 3D environment, e.g. traction forces generated by large cell aggregates over long distances, up to 1.5 mm. Overall, lens-free microscopy techniques favor ease of use and label-free experimentations as well as time-lapse acquisitions of large datasets. Importantly, we consider that these lens-free microscopy technique can thus expand the repertoire of phenomena that can be studied within 2D and 3D organoids cultures. |
