Saturday, September 18th, 2021

Mouse brains seen in unprecedented 3D detail, thanks to new staining technique

Scientists imaged a mouse brain with a new staining method that can image up to four molecular targets at once (three targets shown above).

RIKEN/Creative Commons

Scientists studying brains and other organs and cancerous tumors have long tried to get detailed 3D views of their insides—down to the level of blood vessel and cell type. But producing such images is time-consuming and difficult. Now, dramatic improvements to a 3D imaging technique can reveal the internal components of entire organs or even animals in a simple procedure, researchers report this week. The new tissue staining protocol allows cellular level analyses in unprecedented detail; it could aid research efforts in neuroscience, developmental and evolutionary biology, and immunology, and it could prove useful in diagnosing some cancers and studying damaged brain tissue after death.

To image biological samples in 3D, researchers basically have two main options: They can slice tissues into thin sections and use computer software to reconstruct the whole sample, or they can render biological tissue transparent using special chemicals, which lets researchers view its interior with an optical microscope. To distinguish different cell types, researchers typically stain tissues by soaking them in a cocktail of dyes and chemicals. But getting staining dyes to penetrate organs and large samples has proved difficult.

To tackle this problem, researchers at the RIKEN Center for Biosystems Dynamics Research identified a gel that closely mimics the physicochemical properties of organs that have undergone the tissue clearing process. Starting with computer simulations and following up with laboratory tests, the team optimized the soaking solution temperature, dye and antibody concentrations, chemical additives, and electrical properties to produce the best staining and imaging results. They then tested their method with more than two dozen commonly used dyes and antibodies on mouse and marmoset brains.

Scans of an entire mouse brain and one hemisphere of a marmoset brain—rendered into 3D using light sheet microscopy—revealed the similarity between the two animals’ neural vascular systems, showing the use of the system for comparative anatomy, the researchers report this week in Nature Communications. They also showed that they could simultaneously stain and image up to four molecular targets in a mouse brain, a feat that “has never been reported before,” says Ludovico Silvestri, of the European Laboratory for Non-linear Spectroscopy, who was not involved in the research.

The team also used its technique to image an entire infant marmoset and a small human brain sample—something that could one day lead to new understandings of solid tumors and neurodegenerative diseases. The team says its approach to optimizing staining can be applied to other techniques to advance the entire field of 3D imaging.

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