All tools you need investigate from the smallest synapse to the largest brain slice
In neuroscience, you need the tools to investigate from the smallest synapse to the largest brain slice. HUYGENS offers you DECONVOLUTION for better resolution and less noise in your images. Uniquely, it conserves quantitative information in your images, enabling you to get true quantitative results. Did you also know that HUYGENS offers a comprehensive toolbox for object analysis and large data processing?
A mouse neuron primary cultured cell was fixed and stained with a mouse alpha-tubulin antibody (conjugated to Alexa 568) and Alexa488-phalloidin (F-actin). Microscopy: Nikon A1 confocal (20x Plan Apo VC; NA 0.75) dry. Deconvolution was done with Huygens Essential. Imaged provided by Dr. Motosuke Tsutsumi, Nikon Imaging Center, Hokkaido University, Japan.
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Protocol: Brain cells and the power of STED deconvolution
- Primary astrocytes were fixed and their membranes were permeabilized. Cytochrome C was labeled with a primary antibody and linked to Alexa-488 secondary antibody. (Alexa 488 is a STED compatible dye that emits in the same channel as the commonly used GFP). Primary astrocytes were fixed and their membranes were permeabilized. Cytochrome C was labeled with a primary antibody and linked to Alexa-488 secondary antibody. (Alexa 488 is a STED compatible dye that emits in the same channel as the commonly used GFP).
- Confocal and CW (non-gated) STED (Leica SP8; 100% STED power) images were recorded sequentially.Confocal and CW (non-gated) STED (Leica SP8; 100% STED power) images were recorded sequentially.
- Available metadata was automatically loaded in from the native file-format. Additionally the following settings were set (for more information see STED-Deconvolution: Available metadata was automatically loaded in from the native file-format. Additionally the following settings were set (for more information see STED-Deconvolution:
- blurring is visible in the raw STED image (bottom left) common to all ungated (CW) STED recordings. STED immunity fraction was set to 5%. Huygens is now informed that the PSF has a broad base and will remove blurring accordingly.
- STED depletion factor was set to 30* STED depletion factor was set to 30
- Images were deconvolved using Huygens Express Deconvolution. Express deconvolution estimates the background and signal-to-noise levels from the images automatically, generating reliable care-free image restoration. Huygens Express used GMLE for STED deconvolution and CMLE for Confocal deconvolution.
Confocal and STED image of primary neurons before and after deconvolution. Image kindly provided by Dr. Asier Ruiz Nunez, Achucarro Basque Center for Neuroscience and Department of Neurosciences,. University of Basque Country (UPV/EHU), Leioa, Spain
Toolbox: restoration and analysis of brain tissueHuygens Light Sheet Fluorescence Microscopy (LSFM) deconvolution option supports a range of light sheets types, such as a Gaussian-profile (incl. multiview and flat field), scanning beam, and scanning Bessel beam and lattice. Large LSFM data sets benefit greatly from Huygens efficient RAM use, robust deconvolution algorithms, and GPU acceleration. Huygens Light Sheet deconvolution involves a spatially variant PSF for correcting PSF differences due to changes in light sheet thickness and depth-dependent spherical aberration. Deconvolution can be combined with multi-view fusion, made extremely easy with Huygens Fuser.
mage of Drosophila brain taken with Zeiss Z1 Light Sheet microscope. This multiview dataset consisting of eight views (one of the raw views is shown) was deconvolved and fused with the Huygens Software. Image kindly provided by Dr. Denis Ressnikoff, University Claude Bernard, Lyon, France.
Raw Light Sheet
Neuroscience Image Gallery
Primary hippocampal neuronal culture from P1 C56Bl/6H mice, grown on glass bottom dishes, imaged live in aqueous medium, expressing CMV-driven mCherry (red) and mouse diacylglycerol lipase (with pAcGFP, green) Images were recorded with 1,4 NA 60x oil immersion objective on a Nikon Ti-E inverted confocal microscope. Image kindly provided by Dr. Barna Dudok MSc., Hungarian Academy of Sciences, Hungary.
This is an image of neurons growing in the tissue culture, acquired with spinning disk confocal and deconvolved with Huygens professional. Image kindly provided by Dr. Pawel Pasierbek, Institute of Molecular Pathology, BioOptics (IMP, IMBA, GMI), Vienna, Austria.
The image shows a Drosophila thorax captured with a Zeiss 880 confocal microscope and deconvolved using Huygens Essential. Respiratory tracheal branches are labeled in various colors using stochastic expression of spaghetti monster GFP with defferent epitope tags. Image kindly provided by Prof. Stefan Luschnig, WWU Muenster, Institute for Neurobiology, Germany.
Inhibitory terminals on rat spinal cord neuron. Image shows a pattern of Glycine receptor (red), Gephyrin (blue) and GABA(A) receptor (green) immunolabeling in lumbar L3 segment. Sections were examined with confocal microscopy (Zeiss 780 using 40x obj.), and deconvolved with Huygens Professional. Image kindly provided by Ms. Anna Maciejewska, Laboratory of Reinnervation Processes, Nencki Institute of Experimental Biology, Poland.
Huygens deconvolved and MIP rendered epifluorescence 3D data. Aquaporin 4 (red) shows individual astrocytic end-feet along a blood vessel in a mouse brain part. Thanks to deconvolution, GFAP (green) can be colocalized with AQ4 in small areas, but end-feet and arms of astrocytes can be separately identified. Image kindly provided by Matthew Mitschelen, Dep.of Geriatric Medicine, University of Oklahoma Health Sciences Center, United States.