LAUSR

In the last decade, light sheet fluorescence illumination microscopy (LSFM) became a more and more popular tool for three-dimensional imaging of living organisms and samples. Thanks to the confined excitation volume, this technique provides an elegant solution to combine the optical sectioning capabilities with a limited photo-damage of the sample. For these reasons in the last few years, this fluorescence method became the election tool to study biological processes in living animals and organisms, finding application especially in the field of developmental biology. LSFM has the advantage to combine the optical sectioning of confocal miscopy with the imaging speed typical of the camera-based detection systems. Within this scenario, several are the implementations currently available for LSFM and particular attention has to be addressed to the choice of the proper light sheet architecture able to best suit the requirements of the biological application (review by Zagato et al., 2018). In fact, samples of interest range from cell cultures to tissues and organoids. Within this context, the combination of planar illumination with optofluidic devices provides a tool for high throughput analysis of organoids. Still, the need of developing methods able to image thick biological samples with higher and higher temporal and spatial resolution is without any doubt an important topic that represents one of the current aims in the field. Within this framework, particular attention is addressed to light sheet illumination strategies oriented to increase the axial resolution of light sheet microscopy over large areas. In particular, electrically tunable lens were proven to be an optimal solution to improve axial resolution maintaining the minimal light sheet thickness over a wide field of view (Hedde & Gratton, 2016). Furthermore, methods able to improve axial resolution and simultaneously expand the field of view through phase modulation, obtained by modulating the radially polarized beam with annular binary phase mask (Nhu, Wang, Liu, Kuang, & Liu, 2018). Furthermore, the optimization of Ultramicroscopy, thanks to the use of aspherical optical elements that are able to manipulate the phase and related factors, represents a step forward in the resolution improvement when tissue imaging is performed (Saghafi et al., 2016). This approach shows superior 3D imaging capabilities in tissues when combined with optical clearing methods. In its first implementation, the performances of fluorescence selective plane illumination microscopy were originally demonstrated on embryos and whole animals and it became first popular in developmental biology. Still, beside the imaging capabilities of LSFM in biological samples, different original applications can be proposed. Within this context, it has been demonstrated that light-sheets based lithography technique offer a fast and single-shot process to generate micro-fluidic channels (Mohan & Mondal, 2017). Indeed, light sheet illumination can be implemented for laser interference lithography, thus smoothing the way to the fabrication of periodic microfluidic channels and the generation of complex microstructures.