The Cleared Tissue LightSheet Microscope is 3i’s answer to the question, how to best handle, image, and extract as much valuable information as possible, as quickly as possible, from cleared tissue.
3i Cleared Tissue LightSheet Microscope. (Source: 3i)
Background
Tissue Clearing
Cleared tissue is tissue that has been rendered optically transparent via a series of chemical steps. There are several different procedures for clearing tissue (e.g., CLARITY, iDISCO), each using different chemicals and processes to clear the tissue. These procedures can only be performed on fixed specimens. The goal of all tissue clearing procedures is to change the optical properties of the specimens so that clear, deeper imaging can be performed and large 3d images obtained.
Cleared tissue is ideal for investigating microscopic objects that extend over macroscopic distances. Some examples of these systems include the nervous system, vascular networks, embryonic development, cancer treatments, cell tracking, and many other applications. Combined with an appropriate microscope system, one can image 3d structures within whole organs (e.g., mouse brain), whole organoids, embryos, thick tissue sections, and even whole adult mice and young rats.
If you would like to learn more about tissue clearing, see the journal article, A beginner’s guide to tissue clearing.
Light Sheet Microscopy
Light sheet microscopy is a microscopy technique that quickly images large samples. This is achieved by illuminating the sample with a planar sheet of light or with a scanned beam that approximates a planar sheet. Unlike the point scanning beam of a confocal microscope, a planar sheet of light illuminates large sections of the sample at once. Additionally unlike confocal microscopy, light sheet microscopy decouples the illumination and detection optical pathways. Given these aspects, light sheet microscopy system can be designed to optimize illumination and excitation independently, leading to a higher image frame rate and longer image duration.
If you would like to learn more about light sheet microscopy, see Nikon’s MicroscopyU article.
Cleared Tissue LightSheet Microscope
Cleared tissue and light sheet microscopy seem like a perfect match. However, integration of the two techniques and obtaining consistent, high-quality images can be difficult. 3i’s Cleared Tissue LightSheet (CTLS) microscope was designed to ease the integration of these two techniques. It was also designed to maximize flexibility, allowing for tunable frame rates and resolutions for a wide variety of applications.
Recently, I attended a webinar demonstrating the CTLS microscope. During the demonstration, 3i showed beautiful images of cellular networks in a mouse kidney, an intact mouse femur, whole mouse brain, among other 3d images. The images below were taken from the 3i CTLS website.
Mouse kidney. (Source: 3i)
Mouse femur. (Source: 3i)
The CTLS microscope system can create detailed 3d images of whole organs and large-scale tissue structure. They claimed an entire mouse brain (i.e., tissue 1 cm cubed) can be imaged in 2 hours. During the demonstration, 3i imaged the hindbrain of a mouse brain in about 10 minutes. Within that image, I could see clear individual cells. However, depending on the specimen size and desired resolution, imaging times could be 6-8 hours and up to overnight.
Mouse brain. (Source: 3i)
The CTLS system is built on a Zeiss AxioZoom microscope, consisting of the following key components: a spatial light modulator, a temperature-controlled chamber, a dipping cap for collection objective, and two illumination objectives. The spatial light modulator creates the scanning beam that approximates the planar light sheet. The temperature-controlled chamber and dipping cap maintain constant conditions to facilitate acquisition of high-quality images over long durations. The two illumination objectives allow the system to image the samples from both sides leading to clearer and more intense images.
For more information about 3i’s CTLS microscope, see their website.
The Company, 3i
The small biotechnology company, Intelligent Imaging Innovations (3i), has the aim to “provide advanced multi-dimensional microscopy platforms that are intuitive to use, modular in design, and meet the evolving needs of investigators in the biological research community.” They are based out of Denver, CO. They design and manufacture “technologies for living cell, live cell, and intravital fluorescence microscopy including digital holography, spinning disk confocal, multi-photon, and lightsheet.” They also have their own microscopy software called SlideBook, which is used to control instruments, capture images, and process and analyze data.
For more information on 3i, see their website.
Other Light Sheet Microscopes
The following companies also have light sheet microscopes available:
Miltenyi Biotech Ultramicroscope II
Luxendo light sheet microscopes
Phaseview alpha3 light sheet microscope
CMIF Capabilities
As of the writting of this post, the CMIF does not have a light sheet microscope. If you are interested in tissue clearing or light sheet microscopy, please fill out the form below.
References
“Cleared tissue lightsheet.” Intelligent-imaging.com. 3i, Web. 9 Apr. 2020.
Ariel, P. 2017. “A beginner’s guide to tissue clearing.” International Journal of Biochemistry and Cell Biology, vol. 84. Elsevier Ltd, pp. 35-39.
Allen, J. R. “Light sheet fluorescence microscopy.” MicroscopyU.com. Nikon, Web. 9 Apr. 2020.