title: 4D (x-y-z-t) imaging of thick biological samples by means of Two-Photon inverted Selective Plane Illumination Microscopy (2PE-iSPIM).
abstract: In the last decade light sheet fluorescence microscopy techniques, such as selective plane illumination microscopy (SPIM), has become a well established method for developmental biology. However, conventional SPIM architectures hardly permit imaging of certain tissues since the common sample mounting procedure, based on gel embedding, could interfere with the sample morphology. In this work we propose an inverted selective plane microscopy system (iSPIM), based on non-linear excitation, suitable for 3D tissue imaging. First, the iSPIM architecture provides flexibility on the sample mounting, getting rid of the gel-based mounting typical of conventional SPIM, permitting 3D imaging of hippocampal slices from mouse brain. Moreover, all the advantages brought by two photon excitation (2PE) in terms of reduction of scattering effects and contrast improvement are exploited, demonstrating an improved image quality and contrast compared to single photon excitation. The system proposed represents an optimal platform for tissue imaging and it smooths the way to the applicability of light sheet microscopy to a wider range of samples including those that have to be mounted on non-transparent surfaces.
authors.affiliation: Department of Nanophysics, Istituto Italiano di Tecnologia, Genova, Italy.
authors.forename: Zeno
authors.initials: Z
authors.lastname: Lavagnino
authors.affiliation: Department of Nanophysics, Istituto Italiano di Tecnologia, Genova, Italy.|DIBRIS, Università di Genova, Genova, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Sancataldo
authors.affiliation: Department of Nanophysics, Istituto Italiano di Tecnologia, Genova, Italy.
authors.forename: Marta
authors.initials: M
authors.lastname: d'Amora
authors.affiliation: Department of Neuroscience and Brain Technologies, Istituto Italiano di Tecnologia, Genova, Italy.
authors.forename: Philipp
authors.initials: P
authors.lastname: Follert
authors.affiliation: Department of Neuroscience and Brain Technologies, Istituto Italiano di Tecnologia, Genova, Italy.
authors.forename: Davide
authors.initials: D
authors.lastname: De Pietri Tonelli
authors.affiliation: Department of Nanophysics, Istituto Italiano di Tecnologia, Genova, Italy.|Nikon imaging Center, Istituto Italiano di Tecnologia, Genova, Italy.|Department of Physics, Università di Genova, Genova, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Diaspro
authors.affiliation: Department of Nanophysics, Istituto Italiano di Tecnologia, Genova, Italy.|Nikon imaging Center, Istituto Italiano di Tecnologia, Genova, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Cella Zanacchi