A new HYBRiD technique to make tissue transparent could speed up the study of many diseases

This HYBRiD visualization of a whole mouse chest after SARS-CoV-2 infection shows viral protein in red and tissue structures (lung, blood vessel, bone) in blue. Credit: Scripps Research

The Scripps research technique facilitates the analysis of body-wide biological processes and diseases such as[{” attribute=””>COVID-19 infection.

Scientists at Scripps Research have unveiled a new tissue-clearing method for rendering large biological samples transparent. The method makes it easier than ever for scientists to visualize and study healthy and disease-related biological processes occurring across multiple organ systems.

Described in a paper in Nature Methods on March 28, 2022, and dubbed HYBRiD, the new method combines elements of the two main prior approaches to tissue-clearing technology, and should be more practical and scalable than either for large-sample applications.

“This is a simple and universal tissue-clearing technique for studies of large body parts or even entire animals,” says study senior author Li Ye, PhD, assistant professor of neuroscience at Scripps Research.

Tissue-clearing involves the use of solvents to remove molecules that make tissue opaque (such as fat), rendering the tissue optically transparent—while keeping most proteins and structures in place. Scientists commonly use genetically encoded or antibody-linked fluorescent beacons to mark active genes or other molecules of interest in a lab animal, and tissue-clearing in principle allows these beacons to be imaged all at once across the entire animal.

Learn how a new technique from Scripps Research is making it easier to analyze body-wide biological processes and diseases such as COVID-19 infection. Credit: Scripps Research

Scientists began developing tissue cleaning methods around 15 years ago, primarily with the goal of tracing nerve connections in whole brains. Although the methods work well for the brain, they don’t work so well when applied to other parts of the body or whole bodies, which contain structures that are more difficult to dissolve.

These methods have hitherto used either organic solvents or water-based solvents. The former generally act faster and more powerfully but tend to decrease fluorescent signals. Methods using water-based solvents are more effective at preserving fluorescence, but are impractical for removing non-brain tissue. Additionally, both types of methods require cumbersome and labor-intensive procedures, often using hazardous chemicals.

“An ordinary laboratory usually cannot use these methods routinely and on a large scale,” says Yu Wang, a graduate student at Ye Lab and co-first author of the paper.

The new method devised by Ye and his team uses a sequential combination of organic solvents and water-based detergents, and uses water-based hydrogels to protect tissue molecules that need to be preserved. It often does not require pumping solvents through the sample.

“In many cases, you can just put the whole thing in a jar and keep it in a shaker on your bench until it’s done,” says co-first author Victoria Nudell, a research assistant at Ye Lab. “That makes it convenient and scalable enough for routine use.”

The researchers demonstrated the ease and usefulness of their new method in a variety of applications. These included collaboration with the lab of John Teijaro, PhD, associate professor of immunology and microbiology, to image SARS-CoV-2– infected cells in the entire thorax of mice for the first time, a procedure whose simplicity, with the new method, made it possible to be carried out in a high-level biosafety facility where access to equipment is strictly limited.

Ye and his team are now working with their scientific collaborators on multiple applications of the new method, including tracing nerve pathways in the body.

Reference: “HYBRiD: Hydrogel-reinforced DISCO for clearing mammalian bodies” by Victoria Nudell, Yu Wang, Zhengyuan Pang, Neeraj K. Lal, Min Huang, Namir Shaabani, Wesam Kanim, John Teijaro, Anton Maximov and Li Ye, March 28, 2022 , Natural methods.
DOI: 10.1038/s41592-022-01427-0

“HYBRiD: hydrogel-reinforced DISCO for clearing mammalian bodies” was co-authored by first authors Victoria Nudell and Yu Wang, and by Zhengyuan Pang, Neeraj Lal, Min Huang, Namir Shaabani, Wesam Kanim, John Teijaro, Anton Maximov and Li Yes, all Scripps Research during the study.

The research was supported by the National Institutes of Health (DP2DK128800, K01DK114165), the Dana Foundation and the Baxter Foundation.

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