As an immunologist you need tools that work for you. Huygens understands this and provides easy and reliable deconvolution and analysis tools that help improve your scientific impact. Explore the benefits of deconvolution on our cell biology page or push your resolution to the next level on our super-res page. Check out below recipe for quantitative imaging of viral particles.
Image of a T lymphocyte crawling taken with a Leica SP8 g-STED. The image is deconvolved with Huygens and the final visual effect was obtained using the Huygens surface rendering tool. Blue shows labelled plasma membrane with a specific lipophilic dye, red represents the actin filaments, and yellow internal cytosolic vesicles.
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Protocol: Noisy confocal data restorationConfocal setups allow the user to tune their resolution by setting the pinhole size. A smaller pinhole results in a better resolution, but at the same time increases noisiness because the smaller pinhole transmits less photons for imaging. There are other reasons why your image can be noisy as well: short acquisition times to reduce bleaching or to capture cell movement. HUYGENS DECONVOLUTION is extremely good at restoring noisy images and works synergistically with small pinhole imaging because after deconvolution the image is both denoised and has high-resolution.
- The data file (here a .lif file) is loaded into Huygens. All image acquisition settings are automatically and correctly read-in using Huygens Full File reader
- The image is deconvolved using the default Classic CMLE deconvolution algorithm and standard for background and signal to noise level. If speed is an issue, Good’s Roughness GMLE can be used instead. Check out our deconvolution algorithms for more information.
- Visualise your data in one of our 2D or 3D viewers. Here the image is displayed using a Maximum Intensity Projection (MIP).
The image is a maximum intensity projection (MIP) of a z-stack through a Drosophila macrophage imaged on a Leica TCS SP8X scanning confocal microscope. Objective NA 1.4. Pinhole size 405nm 0.17 AU, 488nm 0.15 AU, 568nm 0.15 AU for obtaining the best possible resolution. Nuclei are DAPI stained (cyan), alpha-tubulin was detected with a primary mouse DM1A antibody and a secondary anti-mouse antibody coupled to Alexa Fluor 488 (magenta), actin was labelled with phalloidin coupled to Alexa Fluor 568 (green). Deconvolution was done with CMLE using Huygens Essential default settings.
Immunology Image Gallery
7-color Cross section image of a Rhesus Macaque lymph node (about 2.0mm X 4.5mm in dimension) showing CD3 (blue), CD4(green), CD8(yellow), PD-1(white), CD20(red), CD56(magenta) and Ki67(cyan). Confocal fluorescent images were acquired using a Leica SP8 microscope (40X/1.30 oil lens) and deconvolved with Huygens software. Image kindly provided by Weiming Yu Ph.D. Laboratory of Systems Biology, NIAID/NIH, United States.
T lymphocyte cells labelled with a lipid marker staining both the cell membrane and cellular comportments (yellow) and a SNAP Tag labelling a membrane protein (green). Confocal z-stack recorded on a Leica SP8 gSTED microscope and deconvolved using Huygens. Visualised using the Huygens surface renderer. Image kindly provided by Dr. Jorge Bernardino de la Serna, Weatherall Institute of Molecular Medicine, University of Oxford, United Kingdom.