Discover ZEISS Lightsheet Z.1, the benchmark for light sheet fluorescence microscopy of in vivo and cleared specimens
Tissue clearing allows you to image deep into highly scattering biological samples (e.g. brains, embryos, organs, spheroids). Tissue clearing is a chemical procedure which replaces light-scattering structures in the sample by molecules which have the same refractive index as the surrounding sample material. This results in enhanced optical penetration to capture fluorescent signals inside of tissues almost without any artifacts caused by scatter. Clearing methods are a promising technique when, for example, investigating neuronal networks in mouse brains with light sheet fluorescence microscopy (LSFM). Our article details the process of imaging cleared tissue with ZEISS Lightsheet Z.1 and gives you all the necessary resources to get started!
ZEISS Lightsheet Z.1 combines the advantages of clearing with light sheet fluorescence microscopy. It lets you image large cleared specimens with exceptional light efficiency, speed and next to no photo damage. Now you can acquire multiple tiles of Z-stacks with several thousand high quality images. A typical imaging speed of 10-40 frames per second reduces your imaging time from hours to minutes.
The sample holder with its easy-to-access interface gives you the flexibility to adapt the sample mounting to your specific needs. Different adapters will either support your sample from below or let you mount the sample hanging from above: you are always free to choose the perfect viewing angle.
With the recently released new optics for clearing applications, ZEISS Lightsheet Z.1 is able to deal with a variety of clearing methods ranging covering refractive index ranges from nd= 1,35 up to nd = 1,48. For example use Lightsheet Z.1 with Clr Plan-Apochromat 20x/1.0 Corr nd=1.38 (± 0,03) to perform imaging experiments on tissue cleared with Scale medium, which has a refractive index of n=1.38. If your aqueous clearing medium has a refractive index of nd=1.45 such as CLARITY, you can choose either Clr Plan-Neofluar 20x/1.0 Corr nd=1.45 (± 0,03) or Lightsheet Z.1 Detection Optics 5x/0.16 to investigate your sample.
Whole mouse embryo, cleared with CUBIC reagent (Susaki et al, Cell, 2014) and imaged with ZEISS Lightsheet Z.1 adapted for cleared tissue imaging. Single-channel fluorescence imaging with glutamic acid decarboxylase-green fluorescence protein (GAD67-GFP) knock-in mouse line. Sample courtesy of Fatima Girach and Tibor Harkany, Medical University of Vienna / Department of Molecular Neurosciences. 5x objective, 54 positions, 600 slices, 170 GB data volume. Rendered with Arivis Vision 4D.
The Center for Brain Research (CBR) at Medical University of Vienna features the first Lightsheet Z.1 system installed in Austria that is equipped for imaging of living and cleared specimens. During a 2-day training course on clearing applications for light sheet microscopy in the Microscopy Labs Munich, Fatima Girach who is a researcher at the CBR generated the stunning dataset of a cleared mouse embryo as part of a tile acquisition exercise. The acquisition was running over lunch break (ca. 30 min) and resulted in this impressive 3D reconstruction.
A recent review by Doug Richardson and Jeff Lichtman from the Harvard Center for Biological Imaging (HCBI) gives an excellent overview of the available clearing methods. The HCBI has been working closely with ZEISS Microscopy to assist in the development of a modified version of Lightsheet Z.1 that is capable of rapidly imaging cleared tissue, while Arivis AG has presented methods for visualization and analysis of the huge imaging data sets (recognized by a Microsoft Life Sciences Innovation Award in 2014).
- Cleared tissue imaging at the Harvard Center for Biological Imaging
- Review: Clarifying Tissue Clearing (Richardson&Lichtman, Cell 2015)
- CLARITY Resources
- SeeDB Resources
- CUBIC (Tainaka et al, Cell 2014)
- PACT (Yang et al, Cell 2014)
- Scale (Hama et al, Nature 2011)
- FRUIT (Hou et al, Frontiers in Neuroanatomy 2015)
ZEISS Application Notes & Quick Guides:
- Application Note: Adjusting Refractive Index for Clearing Applications
- Quick Guide: Sample Holder Universal for Capillaries, Clearing and Large Samples
- Quick Guide: Imaging with Water or Clearing Immersion
- Quick Guide: Tile Scanning
Watch our webinar recording for an overview of in vivo and cleared tissue imaging with ZEISS Lightsheet Z.1!
Visit our Lightsheet Z.1 product page for more application videos, details, and free downloads! If you have questions about a specific application, just use the contact form of the product website to get in contact.