Dataset Information

PiezoGRIN: A High-Pressure Chamber Incorporating GRIN Lenses for High-Resolution 3D-Microscopy of living Cells and Tissues.

ABSTRACT: A high-pressure optical chamber, PiezoGRIN, that facilitates label-free 3D high-resolution live-cell multiphoton microscopy in thick tissue samples is presented. A set of two Gradient Index (GRIN) rod lenses is integrated into the chamber as an optical guide and allows for the adjustment of the focal plane through the sample providing a field of view volume of 450 × 450 × 500 µm (x, y, z). An optical lateral resolution of 0.8 µm is achieved by using two-photon excitation with 150 fs pulses of a 810 nm titanium-sapphire laser at hydrostatic pressures up to 200 MPa. With the PiezoGRIN setup, it is possible to follow pressure-induced changes in subcellular structure of unstained vital mouse skeletal muscle tissue up to 200 µm below the tissue surface.

SUBMITTER: Schneidereit D

PROVIDER: S-EPMC6382305 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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PiezoGRIN: A High-Pressure Chamber Incorporating GRIN Lenses for High-Resolution 3D-Microscopy of living Cells and Tissues.

Schneidereit Dominik D Schürmann Sebastian S Friedrich Oliver O Friedrich Oliver O

Advanced science (Weinheim, Baden-Wurttemberg, Germany) 20181214 4

A high-pressure optical chamber, PiezoGRIN, that facilitates label-free 3D high-resolution live-cell multiphoton microscopy in thick tissue samples is presented. A set of two Gradient Index (GRIN) rod lenses is integrated into the chamber as an optical guide and allows for the adjustment of the focal plane through the sample providing a field of view volume of 450 × 450 × 500 µm (x, y, z). An optical lateral resolution of 0.8 µm is achieved by using two-photon excitat ...[more]

PMID: 30828527

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Project description:Cellular communication constitutes a fundamental mechanism of life, for instance by permitting transfer of information through synapses in the nervous system and by leading to activation of cells during the course of immune responses. Monitoring cell-cell interactions within living adult organisms is crucial in order to draw conclusions on their behavior with respect to the fate of cells, tissues and organs. Until now, there is no technology available that enables dynamic imaging deep within the tissue of living adult organisms at sub-cellular resolution, i.e. detection at the level of few protein molecules. Here we present a novel approach called multi-beam striped-illumination which applies for the first time the principle and advantages of structured-illumination, spatial modulation of the excitation pattern, to laser-scanning-microscopy. We use this approach in two-photon-microscopy--the most adequate optical deep-tissue imaging-technique. As compared to standard two-photon-microscopy, it achieves significant contrast enhancement and up to 3-fold improved axial resolution (optical sectioning) while photobleaching, photodamage and acquisition speed are similar. Its imaging depth is comparable to multifocal two-photon-microscopy and only slightly less than in standard single-beam two-photon-microscopy. Precisely, our studies within mouse lymph nodes demonstrated 216% improved axial and 23% improved lateral resolutions at a depth of 80 µm below the surface. Thus, we are for the first time able to visualize the dynamic interactions between B cells and immune complex deposits on follicular dendritic cells within germinal centers (GCs) of live mice. These interactions play a decisive role in the process of clonal selection, leading to affinity maturation of the humoral immune response. This novel high-resolution intravital microscopy method has a huge potential for numerous applications in neurosciences, immunology, cancer research and developmental biology. Moreover, our striped-illumination approach is able to improve the resolution of any laser-scanning-microscope, including confocal microscopes, by simply choosing an appropriate detector.

Dataset Information

PiezoGRIN: A High-Pressure Chamber Incorporating GRIN Lenses for High-Resolution 3D-Microscopy of living Cells and Tissues.

Publications

<i>PiezoGRIN</i>: A High-Pressure Chamber Incorporating GRIN Lenses for High-Resolution 3D-Microscopy of living Cells and Tissues.

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