Hyperspectral light sheet microscopy.
Ontology highlight
ABSTRACT:
SUBMITTER: Jahr W
PROVIDER: S-EPMC4569691 | biostudies-literature | 2015 Sep
REPOSITORIES: biostudies-literature
Publications
Hyperspectral light sheet microscopy.
Jahr Wiebke W Schmid Benjamin B Schmied Christopher C Fahrbach Florian O FO Huisken Jan J
Nature communications 20150902
To study the development and interactions of cells and tissues, multiple fluorescent markers need to be imaged efficiently in a single living organism. Instead of acquiring individual colours sequentially with filters, we created a platform based on line-scanning light sheet microscopy to record the entire spectrum for each pixel in a three-dimensional volume. We evaluated data sets with varying spectral sampling and determined the optimal channel width to be around 5 nm. With the help of these ...[more]
Similar Datasets
Project description:Hyperspectral microscopy is an imaging technique that provides spectroscopic information with high spatial resolution. When applied in the relevant wavelength region, such as in the infrared (IR), it can reveal a rich spectral fingerprint across different regions of a sample. Challenges associated with low efficiency and high cost of IR light sources and detector arrays have limited its broad adoption. Here, we introduce a new approach to IR hyperspectral microscopy, where the IR spectral map is obtained with off-the-shelf components built for visible light. The method is based on the nonlinear interference of correlated photons generated via parametric down-conversion. In this proof-of-concept we demonstrate the chemical mapping of a patterned sample, where different areas have distinctive IR spectroscopic fingerprints. The method provides a wide field of view, fast readout, and negligible heat delivered to the sample, which opens prospects for its further development for applications in material and biological studies.
| S-EPMC7608807 | biostudies-literature
Project description:Selective-plane illumination microscopy has proven to be a powerful imaging technique due to its unsurpassed acquisition speed and gentle optical sectioning. However, even in the case of multiview imaging techniques that illuminate and image the sample from multiple directions, light scattering inside tissues often severely impairs image contrast. Here we combine multiview light-sheet imaging with electronic confocal slit detection implemented on modern camera sensors. In addition to improved imaging quality, the electronic confocal slit detection doubles the acquisition speed in multiview setups with two opposing illumination directions allowing simultaneous dual-sided illumination. Confocal multiview light-sheet microscopy eliminates the need for specimen-specific data fusion algorithms, streamlines image post-processing, easing data handling and storage.
| S-EPMC4674777 | biostudies-literature
Project description:We developed a deconvolution software for light sheet microscopy that uses a theoretical point spread function, which we derived from a model of image formation in a light sheet microscope. We show that this approach provides excellent blur reduction and enhancement of fine image details for image stacks recorded with low magnification objectives of relatively high NA and high field numbers as e.g. 2x NA 0.14 FN 22, or 4x NA 0.28 FN 22. For these objectives, which are widely used in light sheet microscopy, sufficiently resolved point spread functions that are suitable for deconvolution are difficult to measure and the results obtained by common deconvolution software developed for confocal microscopy are usually poor. We demonstrate that the deconvolutions computed using our point spread function model are equivalent to those obtained using a measured point spread function for a 10x objective with NA 0.3 and for a 20x objective with NA 0.45.
| S-EPMC6879637 | biostudies-literature
Project description:Minimally-invasive optical imaging requires that light is delivered efficiently to limit the detrimental impact of photodamage on delicate biological systems. Light sheet microscopy represents the exemplar in tissue specific optical imaging of small and mesoscopic samples alike. However, further gains towards gentler imaging require a more selective imaging strategy to limit exposure to multiple yet discrete tissues without overexposing the sample, particularly where the information content is sparse or particularly optically sensitive tissues are present. The development of sample-adaptive imaging techniques is crucial in pursuit of the next generation of smart, autonomous microscopes. Herein, we report a microscope capable of performing 4D (x, y, z, t) light patterning to selectively illuminate multiple, rapidly reconfigurable regions of interest while maintaining the rapid imaging speed and high contrast associated with light sheet microscopy. We illustrate this utility in living zebrafish larvae and phantom samples.
| S-EPMC6018501 | biostudies-literature
Project description:Light-sheet fluorescence microscopy (LSFM) has been present in cell biology laboratories for quite some time, mainly as custom-made systems, with imaging applications ranging from single cells (in the micrometer scale) to small organisms (in the millimeter scale). Such microscopes distinguish themselves for having very low phototoxicity levels and high spatial and temporal resolution, properties that make them ideal for a large range of applications. These include the study of cellular dynamics, in particular cellular motion which is essential to processes such as tumor metastasis and tissue development. Experimental setups make extensive use of microdevices (bioMEMS) that provide better control over the substrate environment than traditional cell culture experiments. For example, to mimic in vivo conditions, experiment biochemical dynamics, and trap, move or count cells. Microdevices provide a higher degree of empirical complexity but, so far, most have been designed to be imaged through wide-field or confocal microscopes. Nonetheless, the properties of LSFM render it ideal for 3D characterization of active cells. When working with microdevices, confocal microscopy is more widespread than LSFM even though it suffers from higher phototoxicity and slower acquisition speeds. It is sometimes possible to illuminate with a light-sheet microdevices designed for confocal microscopes. However, these bioMEMS must be redesigned to exploit the full potential of LSFM and image more frequently on a wider scale phenomena such as motion, traction, differentiation, and diffusion of molecules. The use of microdevices for LSFM has extended beyond cell tracking studies into experiments regarding cytometry, spheroid cultures and lab-on-a-chip automation. Due to light-sheet microscopy being in its early stages, a setup of these characteristics demands some degree of optical expertise; and designing three-dimensional microdevices requires facilities, ingenuity, and experience in microfabrication. In this paper, we explore different approaches where light-sheet microscopy can achieve single-cell and subcellular resolution within microdevices, and provide a few pointers on how these experiments may be improved.
| S-EPMC6362405 | biostudies-literature
Project description:Light sheet microscopy in the museumLight sheet microscopy (or selective plane illumination microscopy) is an important imaging technique in the life sciences. At the same time, this technique is also ideally suited for community outreach projects, because it produces visually appealing, highly dynamic images of living organisms and its working principle can be understood with basic optics knowledge. Still, the underlying concepts are widely unknown to the non-scientific public. On the occasion of the UNESCO International Year of Light, a technical museum in Dresden, Germany, launched a special, interactive exhibition. We built a fully functional, educational selective plane illumination microscope (eduSPIM) to demonstrate how developments in microscopy promote discoveries in biology.Design principles of an educational light sheet microscopeTo maximize educational impact, we radically reduced a standard light sheet microscope to its essential components without compromising functionality and incorporated stringent safety concepts beyond those needed in the lab. Our eduSPIM system features one illumination and one detection path and a sealed sample chamber. We image fixed zebrafish embryos with fluorescent vasculature, because the structure is meaningful to laymen and visualises the optical principles of light sheet microscopy. Via a simplified interface, visitors acquire fluorescence and transmission data simultaneously.The eduspim design is tailored easily to fit numerous applicationsThe universal concepts presented here may also apply to other scientific approaches that are communicated to laymen in interactive settings. The specific eduSPIM design is adapted easily for various outreach and teaching activities. eduSPIM may even prove useful for labs needing a simple SPIM. A detailed parts list and schematics to rebuild eduSPIM are provided.
| S-EPMC4999180 | biostudies-literature
Project description:Contactless sample confinement would enable a whole host of new studies in developmental biology and neuroscience, in particular, when combined with long-term, wide-field optical imaging. To achieve this goal, we demonstrate a contactless acoustic gradient force trap for sample confinement in light sheet microscopy. Our approach allows the integration of real-time environmentally controlled experiments with wide-field low photo-toxic imaging, which we demonstrate on a variety of marine animal embryos and larvae. To illustrate the key advantages of our approach, we provide quantitative data for the dynamic response of the heartbeat of zebrafish larvae to verapamil and norepinephrine, which are known to affect cardiovascular function. Optical flow analysis allows us to explore the cardiac cycle of the zebrafish and determine the changes in contractile volume within the heart. Overcoming the restrictions of sample immobilisation and mounting can open up a broad range of studies, with real-time drug-based assays and biomechanical analyses.
| S-EPMC6368588 | biostudies-literature
Project description:Optical clearing combined with deep imaging of large biological specimen allows organ-wide visualization of cells in three dimensions (3D) to explore regenerative processes in a spatial context. Here, we investigate the dynamics of airway regeneration following toxin-mediated epithelial injury in cleared whole lung preparations by light sheet microscopy. We use a recently developed knock-in mouse strain labeling bronchiolar Club cells (Scgb1a1-mCherry) to define an optimal clearing procedure that efficiently preserves genetically encoded fluorophores. Dehydration in pH-adjusted tert-butanol followed by clearing in ethyl cinnamate maintained maximum mCherry fluorescence while preventing unfavorable background fluorescence. We apply this technique to depict the course of bronchiolar epithelial renewal from an acute injury phase to early and late recovery stages. 3D reconstructions of whole lungs demonstrate near-complete loss of secretory Club cells throughout the entire respiratory tract 3 days post naphthalene (dpn). Multiple foci of regenerating Club cells emerge at 7 dpn, predominantly at airway bifurcations and in distal terminal bronchioles-anatomical regions assumed to harbor distinct stem/progenitor cells subsets. At 21 dpn, clusters of newly formed Club cells have largely expanded, although the bronchiolar epithelial lining continues to regenerate. This study identifies regional stem cell niches as starting points for epithelial recovery, underscores the enormous regenerative capacity of the respiratory epithelium and demonstrates the power of whole lung 3D imaging for evaluating the extent of pulmonary damage and subsequent repair processes.
| S-EPMC7910255 | biostudies-literature
Project description:Microbial colonization of the digestive tract is a crucial event in vertebrate development, required for maturation of host immunity and establishment of normal digestive physiology. Advances in genomic, proteomic, and metabolomic technologies are providing a more detailed picture of the constituents of the intestinal habitat, but these approaches lack the spatial and temporal resolution needed to characterize the assembly and dynamics of microbial communities in this complex environment. We report the use of light sheet microscopy to provide high-resolution imaging of bacterial colonization of the intestine of Danio rerio, the zebrafish. The method allows us to characterize bacterial population dynamics across the entire organ and the behaviors of individual bacterial and host cells throughout the colonization process. The large four-dimensional data sets generated by these imaging approaches require new strategies for image analysis. When integrated with other "omics" data sets, information about the spatial and temporal dynamics of microbial cells within the vertebrate intestine will provide new mechanistic insights into how microbial communities assemble and function within hosts.
| S-EPMC3952068 | biostudies-literature
Project description:Attenuation of optical fields owing to scattering and absorption limits the penetration depth for imaging. Whilst aberration correction may be used, this is difficult to implement over a large field-of-view in heterogeneous tissue. Attenuation-compensation allows tailoring of the maximum lobe of a propagation-invariant light field and promises an increase in depth penetration for imaging. Here we show this promising approach may be implemented in multi-photon (two-photon) light-sheet fluorescence microscopy and, furthermore, can be achieved in a facile manner utilizing a graded neutral density filter, circumventing the need for complex beam shaping apparatus. A "gold standard" system utilizing a spatial light modulator for beam shaping is used to benchmark our implementation. The approach will open up enhanced depth penetration in light-sheet imaging to a wide range of end users.
| S-EPMC7229186 | biostudies-literature